Olivier Emile
email: olivier.emile@univ-rennes1.fr

Janine Emile

Observing wave/particle duality of light using topological charge

Keywords: Light orbital angular momentum, Mach-Zehnder interferometer, Light wave/particle duality

Wave particle duality, also called complementarity, is deeply rooted in the heart of quantum theory. It is fully exemplified in the famous Wheeler's delayed choice experiment where the choice of the wave nature (ability to interfere) or the particle like behavior (path distinguishability) is introduced a posteriori. We perform here a delayed choice experiment in a Mach-Zehnder interferometer, using a classical laser beam and twisted light in a given mode. We entangle the polarization and the twisted internal degrees of freedom, with the which-path-information external degree of freedom of the beam. The particle behavior of light arises from the quantization of the orbital angular momentum. It is demonstrated from torque and light power measurements within 10% accuracy. We then experimentally evidence that the particle or wave behavior of light can be chosen a posteriori, even after the light has left the interferometer, at the moment of the detection.

INTRODUCTION

The complementary discussion is at the very basis of quantum mechanics [Bohr1928, Feynman1963, Scully1991, Bertet2001]. It is best evidenced in the so-called Wheeler's delayed choice experiment [Wheeler1978, Wheeler1984, Durr1998, Legget2009, Ma2016] that was first proposed for electrons. Briefly, as the output beam splitter of a Mach-Zehnder (MZ) interferometer is inserted, electrons in the two paths interfere. They behave as waves. When the beam splitter is removed, electrons travel either in one or the other arm. They behave as particles. Wheeler's claim is that, by delaying the choice for fixing the position of the final beam splitter, forces electrons to somehow decide whether to behave like particles or waves, long after they enter the interferometer [Hiley2006]. This has been experimentally demonstrated in optics using light from a single photon source [Braig2003, Jacques2007, Tang2012] instead of electrons. Like for electrons, when the beam splitter is removed, photons travel either in one arm or in the other arm, and are detected at a single photon level. They behave as particles. When the beam splitter is inserted, photons interfere and behave as wave. Besides, since a long interferometric device is used, the choice can be made even after the light has already entered the interferometer. However, since a single photon source is used, the evidence of the wave nature of photons is not at all immediate. Indeed, the interference is due to a change in the optical path within the interferometer and its observation is constructed photon by photon and depends strongly on the statistics of the source. Moreover, the corpuscular nature of the light from the source has to be distinguished from the quantum nature of the detection.

On the other hand, Electromagnetic (EM) fields carry angular momentum that may be detected via its transfer to matter [Beth1935, Emile2014a]. Within the paraxial approximation, two different forms of angular momentum may be distinguished. The first one is associated with the dynamical rotation of the direction of the fields around the propagation direction (Spin Angular Momentum, SAM), also called circular polarization. The second one is associated with the dynamical rotation of the light rays or the Poynting vector around the beam axis [Molina2007, Barnett2017] (Orbital Angular Momentum, OAM). SAM is an intrinsic concept, that can be defined locally at a given position [Brown2011]. In particular, this means that two orthogonally polarized beams cannot interfere, even if only a small part of the beams is considered. On the contrary, OAM is defined from the whole beam, implying in particular that two OAM orthogonal beams can interfere [Courtial1998, Vickers2008]. This has dramatic consequences in light entanglement experiments for example [Fickler2012]. It may also shine some new light in fundamental tests of quantum theory such as complementarity. Indeed, more specifically, could Wheeler's delayed choice experiment be carried out readily and unambiguously with a standard source, using OAM?

EXPERIMENT

To address this issue, we have developed the experimental set-up described in figure 1. We use here the fundamental mode of a helium-neon laser (Melles Griot, λ = 633 nm, P=1 mW) that is transformed in a higher order Laguerre Gaussian (LG) mode with a spiral phase plate (SPP) [Beijersbergen1994]. The splitting of the beam is performed with a PBS set at 45 • from the polarization of the laser. The two arms of the MZ interferometer have the same intensity and correspond to linear vertical (along y) and horizontal (along x) polarizations of light respectively. The length of the interferometer is z = 30 cm.

The main difference from a usual MZ interferometer here, is that we use a LG laser beam at the entrance of the interferometer. Then, half of the light intensity travels in arm (1), with a polarization along x in a LG mode with a topological charge or OAM = +2. The other half travels in arm (2), with a polarization along y in a LG mode transformed from = +2 to = -2 by two identical cylindrical lenses separated from twice their focal length [Padgett2002].

A second PBS then recombines the two arms that have orthogonal polarizations and conjugated topological charges. We carefully adjust mirrors M 1 and M 2 so that the two recombined beams overlap exactly. Note that the apparently longer length of arm (1), is compensated by the travel length inside the cylindrical glass lenses (CL), and also because the PBS are positioned little off axis, so that the travel path of the beam running through arm (1) is longer in the PBS than for the beam running through arm (2). The interferometer is thus balanced.

We have entangled the which-path information (arm (1) or arm (2)) with the polarization (along x or y) and the topological charge + or -, of the beam. We made them non-separable. At the end of the interferometer the light beam is in a coherent superposition of (i) light that has travelled through arm (1), with a = +2 topological charge and a polarization along x, and (ii) light that has travelled through arm (2), with a = -2 topological charge and a polarization along y. In a more formal way, this coherent superposition of the EM field could be written (without normalization),

|ψ M Z EM = |1 ⊗ | + ⊗ |x + |2 ⊗ | -⊗ |y , (1)
the first item refers to the which-path information, either arm (1) or arm (2), the second item refers to the topological charge (OAM) of the beam and the last one to the polarization state (SAM).

RESULTS

The detection scheme is displayed in figure 2. The light coming from the twisted MZ interferometer of figure 1 crosses a half wave plate (λ/2) that can be rotated and enters a PBS. Light is split in two parts and detected on two screens with the naked eye.

Particle behavior

When the axes of the λ/2 are aligned with the axes of the PBS and correspond to the x and y directions, the light on A is polarized along x, has travelled through arm (1), and has a = +2 topological charge. Conversely, the light on B is polarized along y, has travelled through arm (2), and has a = -2 topological charge. In a more formal writing, the signal on screen A and B is the projection of equation (1) on |x and |y respectively. It writes

|ψ A EM = |1 ⊗ | + ⊗ |x , (2)
and

|ψ B EM = |2 ⊗ | -⊗ |y (3)
where |ψ A EM and |ψ B EM are the states of the light observed on the screens A and B respectively. Thus light that has travelled through arm (1) ends up in A, and light that has travelled through arm (2) ends up in B. It is as if, there was no beam splitter at the output of the interferometer in figure 1 that recombines the beams. What do we detect experimentally?

The light intensities recorded on screens A and B look like the same. They have seemingly a vortex structure that may correspond to twisted OAM beams. We have probed their topological charge with Young's double slit experiment [Emile2014b]. The fringes after the slits correspond to = +2 and = -2 topological charges respectively (see figure 3). The purity of the modes is better then 95%, from the interference pattern. This proves that we have experimentally entangled the which pass information with the polarization and the OAM state of light, as expected from equation (1). On screen A, light has travelled through arm (1) is polarized along x and has a topological charge = +2, in agreement with equation (2). Conversely, on screen B, light has travelled through arm (2) is polarized along y and has a topological charge equal to = -2, in agreement with equation (3). Nevertheless, up to now, the results can be explained classically and do not prove the particle nature of light. Let us couple then the OAM with a macroscopic mechanical property. We have thus measured the available torque on both outputs of the PBS (see figure 4).

The torque detection here is similar to what has been done for SAM [Delannoy2005], or even for OAM in the case of radio emission [Emile2014a, Emile2016]. A 1.5 mm-diameter absorbing black paper (density 180 g.m -2 ) hangs from a-10 cm-long ordinary cotton thread. The whole system is set in a vacuum chamber (pressure of 0.5 Pa). The beam coming out the PBS is focalized on the black paper with a 5-cm focal length ordinary lens. We register the rotation of the paper with a camera during 6 min (see figure 4). We then measure the rotation angle that is in the ten degrees range. Since the rotation angle is small and the thread is long, the restoring torque is negligible. Besides, since the system is in a vacuum chamber, the friction is negligible either. The possible heating of the black paper has no influence on the torque. We observe a uniformly accelerated rotation. We then deduce the angular acceleration γ that is equal to γ = 2.25 ± 0.05 × 10 -6 rad.s -2 . We measure exactly the reversed acceleration for the beam at the other output of the PBS. We evaluate the moment of inertia J of the piece of paper that is equal to J = 4.47±0.09×10 -14 kg.m 2 . We then calculate the torque Γ = Jγ. It equals Γ = 1.00 ± 0.05 × 10 -19 N.m for = +2. The torque is reversed for = -2. Assuming that all the light impinging on the paper is absorbed, due to angular momentum conservation, Γ corresponds to the total OAM carried by the light. Before evacuating the chamber, we measure independently, at the same location the light power P that is equal to P = 0.30 ± 0.015 mW.

The ratio Γω/P equals Γω/P = 2.01 ± 0.15, ω being the light pulsation. Measuring only an integer number demonstrates that the OAM is quantized. We have checked that when we change the value of ( = ±1, ±2, ±3, ±4), the ratio Γω/P still equals . Γ/2 corresponds to the number of particles detected per second. Thus, because of the mechanical detection, independently of the photon statistics, neither on the nature of the light source, nor on the detection quantification, we detect particles carrying 2 OAM for = +2 and -2 OAM for = -2, respectively, at an average rate of N= Γ/2 = 29.5 × 10 14 particles per second. Unambiguously, light behaves as particles. Besides, the detection is performed with the naked eye, independently from any quantization nature consideration of the detector system.

One may think that this result could be obtained classically. Indeed, if one considers a field carrying orbital angular momentum with a topological charge equal to , following a straightforward "classical" calculation, one finds that Γω/P equals . However, doing so, one implicitly considers that the topological charge is quantized. Because of cylindrical symmetry, it can only be equal an integer number. This reasoning here is very similar to what has been previously followed in the case of polarization [Raman1931], but also recently in the study of acoustic waves measuring both radiation pressure and torque [Demore2012].

Actually, from a mechanical point of view [Truesdell1968], angular momentum is a physical observable in its own right, in general independent of and not derivable from energy. The knowledge of one of these quantities doesn't imply the knowledge of the other. Energy on one side and orbital angular momentum on the other side are indeed truly independent quantities.

Wave behavior

Let us rotate the λ/2 by 45 • . The lights arriving on screens A and B are the projections of the light polarization on the eigen directions of the PBS, at 45 • from x and y. On screen A, it is the sum of part of the light that has travelled through arm (1), with a = +2 topological charge and part of the light that has travelled through arm (2), with a = -2 topological charge. In a more formal way, it is the projection of equation ( 1) on (|x + |y )

|ψ A EM = |1 ⊗ | + + |2 ⊗ | -, (4)
and the light on B is the difference of part of the light that has travelled through arm (1), with a = +2 topological charge and part of the light that has travelled through arm (2), with a = -2 topological charge. It is the projection of equation ( 1) on (|x -|y ). It then writes,

|ψ B EM = |1 ⊗ | + -|2 ⊗ | -. (5)
However, as already stated, two conjugated LG modes interfere. Thus light that has travelled in arm (1) interferes with light that has travelled through arm (2). We shall detect interference patterns corresponding to conjugated LG beams [Emile2017]. The light then behaves as waves.

Figure 5 shows experimental pictures of the light intensity distribution detected on the screens A and B. They are 4 petals daisy flowers, as expected from equations (4) and (5) and from the interference of conjugated LG beams. They are observed directly with the naked eye. The interference pattern on screens A and B are at 45 • from each other as expected. This also validates the use of equations (4) and (5) to describe the character of light. Of course the torque measurement in this configuration is equal to zero. The two macroscopic mechanical torques cancel, but light interferes. Finally, this clearly experimentally shows that, with this orientation of the λ/2, light behaves as wave.

DISCUSSION

The choice to insert or remove the output beam splitter is made a posteriori. We have even increased the distance between the output of the interferometer and the detection scheme to d = 15 m, in order to fulfill timing, also called sometimes relativistic separation condition [Jacques2007]. We mean here that the choice of the switching must be space like separated from the entering of light in the interferometer. A fast electro-optical modulator (EOM) replaces the hand-rotated λ/2. It acts as an adjustable birefringent plate. When no voltage is applied, the axes of the PBS are aligned with x and y. This situation corresponds to the removal of the beam splitter at the end of the interferometer in the original Wheeler's proposal. Experimentally, as in figure 4, we measure the torque and the light power. We again find that light behaves as particles.

When a voltage, leading to a rotation of the polarization eigen-states by 45 • , is applied, the axes of the PBS are at 45 • regarded to the polarization of the light in the arms of the interferometer. It is equivalent to the insertion of the beam splitter at the end of the interferometer in the original Wheeler's proposal. Experimentally, as in figure 5, we observe four petals daisy flowers.

We find that light, here also, behaves as waves. The switching time of the EOM can be as fast as 10 ns, corresponding to a 3-meter light propagation. This is much less than d. The switch can be random, thus fulfilling timing separation conditions. Of course, the fast switching is not evidenced with the naked eye. We use three fast photodiodes located on the same radius and rotated by π/10 from one another, regarded to the beam axis. If the three photodiodes measure the same intensity, it corresponds to figure 4, whereas when an imbalanced between the photodiodes is measured, it corresponds to a daisy flower and to figure 5.

Besides, it has to be noted that, contrarily to what has been previously demonstrated [Jacques2007], the choice to insert or remove the output beam splitter is performed not only after the light has entered the interferometer, but even long after the light has left the interferometer and the output beam splitter. It is performed just before the light detection.

Could we even demonstrate at the same time the particle and wave behavior using classical state of light? A similar question has been recently asked with a two-photon quantum source [Peruzzo2012, Kaiser2012]. However, in their experiments, the authors observe an entity that is partially a wave and a particle. Here, we intend to probe the wave behavior and the particle behavior at the same time. We shall then use a single set-up to probe complementarity. Let us modify the detection scheme of figure 2 now depicted in figure 6. Light first crosses an ordinary beam splitter. Part of the light is sent to a PBS which axes are aligned with the polarization of the light in the arms of the interferometer. The detection A or B probes which way of the interferometer the light passes through. It is related to the particle like nature of light. It leads to the same results as in figure 4.

The other part of the light is sent to an other PBS which axes are at 45 • compared with the polarization of the light in the arms of the interferometer. It probes the wave nature of light. It leads to the same result as in figure 5. Thus, at the same time, within the same interferometer, part of the light behaves as particles, whereas, simultaneously, the other part of the neighboring light behaves as wave that interferes.

CONCLUSION

We have performed a Wheeler's type experiment using a classical He-Ne laser and conjugated OAM beams in the arms of the interferometer. The wave behavior arises from the interference of the two paths of the modified interferometer, as in usual MZ interferometer with light. The particle behavior arises from the OAM that is indeed quantized. Moreover, we have answered the question of when the decision whether light will display wave like or particle like behavior [Hiley2006] is made. As recently demonstrated for atoms [Manning2015], the decision here for light is made at the point of measurement. Moreover, this decision is independent from the behavior of the neighboring light at the same time, at the same location, in the same interferometer.

These counterintuitive results may then lead to deny the physical reality of the wavefunction to maintain causality. However, a recent work has directly verified the physical reality of the wavefunction [Zhou2017]. It turns out that our results are in agreement with this recent work since, at the output of the interferometer, the nature of light is a vector vortex state, i.e. the state described by its wavefunction, that collapses at the moment of the measurement.

Acknowledgments. The authors thank Jean René THEBAULT for technical assistance.

FIG. 1 .

 1 FIG. 1. Twisted MZ interferometer design. The fundamental mode of a helium-neon laser is transformed into a = +2 LG mode with a SPP. Its polarization is at 45 • from the axes of the polarizing beam splitter (PBS). The PBS splits the beam in two parts. One part is transformed into a = -2 LG mode thanks to two cylindrical lenses (CL), focal length f = 5 cm. The two beams are then recombined with the help of a second PBS. M1, M2: mirrors.

  FIG. 2. Detection scheme. The light intensity from the MZ interferometer is split in two parts by a PBS, and observed with the naked eye, on two screens A and B. Depending on the orientation of the half wave plate (λ/2), light could behave as wave or particles.

  FIG. 3. Measure of the topological charge of the beams after. The axis of the PBS correspond to the polarization of the light in the arms of the interferometer. Vortex beams are detected. Their topological charges are measured either with Youngs double slit [Emile2014b] or Fresnel Bi-prism [Emile2014c] technics. They are equal to = +2 for arm (1) and = -2 for arm (2) respectively.

[FIG. 4 .

 4 FIG. 4. Particle behavior. As the axes of the PBS correspond to the x and y directions, vortex beams are detected. Their topological charges equal to = +2 and = -2 and are measured by the torque deduced from the rotation of an absorbing disk.

FIG. 5 .FIG. 6 .

 56 FIG. 5. Wave behavior.As the axes of the PBS are at 45 • from the x and y directions, = +2 and = -2 LG beams interfere on both output of the PBS, leading to 4 petals daisy flowers.


A Itié

F Polian

J Z Baudelet

Jiang

F Decremps

J Pellicer-Porres

F Datchi

J P Itié

A Polian

F Baudelet

J Z Jiang

Trapping of cubic ZnO nanocrystallites at ambient conditions

Keywords:

Trapping of cubic ZnO nanocrystallites at ambient conditions

1 With a direct band gap of 3.4 eV at ambient conditions, wurtzitic ZnO (w-ZnO), like GaN or 6H-SiC, is intensively studied in view of potential applicability in the field of optoelectronics (e.g. laser or light-emitting diodes) [START_REF] Kawazoe | [END_REF]. Furthermore, in comparison with its previous competitors, w-ZnO combines several unique electrical, acoustic and chemical properties which make it one of the most technologically relevant binary compounds [2].

Recently, it has been shown that intentionally as well as unintentionally doped w-ZnO give rise to high-quality n-type (electron) conductivity that directly comes with its wurtzite ambient condition stable structure [3]. The p-type w-ZnO conductivity has been theoretically predicted to be increased by doping with nitrogen through the use of NO or NO 2 [4] and some recent publications demonstrated that p-type w-ZnO thin films could be fabricated [5][6][7]. However, physical limitations preclude the synthesis of high p-type (hole) conductivity, and therefore, the practical applicability of novel short-wavelength optoelectronic devices based on p-n junctions [8]. On the other hand, ZnO in its ionic NaCl high pressure phase (c-ZnO) has an electronic structure with an indirect band gap that may be compatible for high p-type doping. The possibility to trap this high pressure phase at ambient condition would therefore be of great interest. Several high pressure structural investigations of ZnO have been done in the last two decades [9] and all agree that the w-ZnO → c-ZnO transition is reversible at ambient temperature (with a transition pressure of 9 GPa upon increasing pressure and 2 GPa upon decreasing pressure [10]). In this work, we demonstrate a way to obtain c-ZnO crystallites metastable at ambient conditions based on the peculiar physical features observed in the nanocrystal regime.

Unlike extended solids, nanocrystals are nearly interior defect-free and externally faceted with a defined shape directly related to the crystallographic structure. Under pressure, a structural transformation in these systems occurs with a single nucleation center per crystallite [11] which is usually associated with a shape change [12,13]. Thus, the average surface tension as well as the surface areas of the nanocrystallites should obviously affect the internal energy difference between the high and low pressure structure, unlike bulk crystals.

For example, measurements on CdSe [11] indicate that both forward and backward pressure transition are particle-size dependents. In a same manner, Jiang [14] found an enhancement of the upstroke transition pressure (15.1 GPa) in 12(2) nm size ZnO nanocrystals compared to the bulk value. After a decompression from 52.5 GPa down to ambient pressure, a coexistence of the w-and c-phases was recovered with a large fraction of NaCl phase. We reproduced this experiment with the same type of nanopowder (12(2) nm) and we found that the c-fraction of recovered sample after decompression at ambient conditions increases with the maximum pressure reached upon compression. This phenomenon can not be ascribed to a size distribution effect, since above the transition pressure the whole sample is observed to be in the high pressure phase independently of the maximum pressure reached (which also rejects a possible effect due to a pressure gradient). From the work of Tolbert [11] and Wickham [13], we suggest that this phenomenon is related to the shape change of the crystallites which occurs at the phase transition. At room temperature, and pressure above the phase transition, a fraction of the crystallites may acquire an unstable shape compared to the equilibrium configuration one (despite the interior high-pressure structure). Since the shape change (due to atomic diffusion) is mainly determined by a dynamic processes, heating at high pressure should help to anneal the unstable shape to the equilibrium one.

Finally, heating would lead to a larger number of rocksalt nanocrystallites of ZnO trapping at ambient conditions.

Obtained from planetary ball mill, nanocrystalline powder of w-ZnO with average size of 12(2) nm was used in this work. The high pressure cell was a membrane diamond anvil cell (DAC) specifically designed for high temperature studies [15]. The DAC was surrounded by a heater and a thermocouple was glued on one end of the diamond anvil to determine the average temperature on the sample with an accuracy of 20 K. A rhenium gasket was preindented from 200 to 30 µm, and a 150 µm hole was drilled in the center by sparkerosion. Silicon-oil was used as a pressure transmitting medium because it is amorphous and chemically inert. Pressure was measured using the fluorescence emission of a ruby ball [16] placed into the gasket hole. The accuracy was better than 1 GPa at the maximum pressure reached.

Combined x-ray diffraction (XRD) and x-ray absorption spectroscopy (XAS) experiments

have been performed at the LURE synchrotron facility in Orsay (France), giving respectively long-range and local structure. The powder x-ray diffraction measurements were made in the energy dispersive mode with the wiggler at DW11 station. After the energy calibration of the Ge detector, the 2θ angle was determined by collecting diffraction patterns of a gold sample placed in between the anvils. The polychromatic x-ray beam was collimated to a 50×50 µm 2 spot centered on to the gasket hole. Absorption spectroscopy was performed at the D11 dispersive XAS station at the Zn K edge (see reference [17] for technical details) and the Zn-O distance was deduced from the experimental data using the FEFF6 [18] codes for theoretical calculations of x-ray absorption fine structure including multiple-scattering pathways.

The sample was first pressurized at ambient temperature up to 15 GPa and then heated to 550 K. Under these conditions, ZnO was in a pure rocksalt structure (F m3m) as expected from the phase diagram of the extended crystal [19]. After cooling down to 300 K and a gradual decompression to ambient pressure, the crystallographic structure of the sample recovered from the previous P-T path was examined in detail by XRD and XAS.

X-ray diffraction spectra recorded before and after high-pressure treatment are reproduced in fig. 1. Exposure times between 30 and 40 min were sufficient to collect diffraction patterns with suitable signal to noise ratio. The five peaks observed in the recovered sample spectrum are unambiguously indexed to a cubic cell and used to compute the structural parameter, leading to a 0 =4.278(3) Å (i.e. V 0 =19.57(1) Å3 ).

The local structure (up to 8 Å) around the Zn atoms of ZnO has been probed by XAS. X-ray absorption fine structure (XAFS) spectra before and after high-pressure hightemperature cycle are compared in fig. 2 with that of pure rocksalt zinc oxide recorded at 15 GPa and 300 K (after cooling down from 550 K). Here again, the recovered sample is found to be in a sixfold coordination scheme (within uncertainty of 5%, given by the crystalline structure sensitivity of both XRD and XAS techniques) corresponding to a NaCl type structure (in w-ZnO, the coordination number is four).

Figure 3 shows the magnitude of the experimental and calculated Fourier transform.

The theoretical data have been fitted for a cubic structure to the experimental spectra in the [0.5-3.65 Å] range with only four parameters: the first-neighbor distance d Zn-O , the corresponding variance σ 2 , the theoretical energy origin E 0 and a nonstructural parameter S 2 0 for amplitude correction. The best fit gives d Zn-O =2.14(1) Å (i.e. V 0 =19.60(3) Å3 ), in very good agreement with the present XRD data, as well as with the zero-pressure extrapolation of high pressure x-ray diffraction data obtained on bulk cubic ZnO: V 0 =19.66 Å3 [19], V 0 =19.48 Å3 [20] or V 0 =19.60 Å3 [9].

In conclusion, while the wurtzite-to-rocksalt phase transition has been shown to be reversible for bulk ZnO crystals, this property is demonstrated to be drastically altered by the reduction of grain size. From the combined XRD and XAS observations, we argue that high-pressure (15 GPa) heat (550 K) treatment of ZnO nanocrystallites yields to a single metastable cubic phase of zinc oxide at room conditions. This result is of most interest since this metastable phase may open a broad range of new technical applications as heterojunctions for photodetection and photovoltaic devices [21]. Moreover, while a small volume of c-ZnO powder has only been obtained in the present work (through the use of diamondanvil cell), the P -T conditions required to obtain this sample can also be reached by multi-anvil high-pressure apparatus where the sample volume is of 100 mm 3 . Finally, and from a more general point of view, the present study indicates that the phase diagram of semiconductors with reduced size may be different from that of the bulk compounds and deserve new studies.

Figure 1 :

 1 Figure 1: X-ray energy dispersive spectra for nanopowder of ZnO at ambient conditions before (bottom pattern) and after (top pattern) the high-pressure high-temperature (15 GPa, 550 K) cycle. A single NaCl phase spectrum of ZnO measured at 10 GPa and 1000 K (V=19.16 Å3 ) is also shown for comparison.

Figure 2 :

 2 Figure 2: Zn K-edge XAFS kχ(k) experimental data for nanopowder of ZnO before (bottom pattern) and after (top pattern) high-pressure high-temperature (15 GPa, 550 K) cycle, compared with the experimental kχ(k) signal of pure c-ZnO recorded at 15 GPa and 300 K.

Figure 3 :

 3 Figure 3: Magnitude of the Fourier transform of the recovered sample k 2 χ(k) spectra (solid line) compared with the best fit of theory (dotted line) including all single and multiple-scattering paths between 0.5 and 3.65 Å. The best fit is obtained with d Zn-O =2.14(1) Å, σ 2 =0.01 Å2 , E 0 =9659.4 eV and S 2 0 =0.83.

1

 1

  FIG.1:

  Figure 3

Acknowledgments

This research has been partly supported by a Marie Curie Fellowship of the European Community Human Potential Program under Contract No. HPMF CT2000-00764.

Disclaimer: The European Commission is not responsible for any views or results expressed.


Nicolas Le Roux
email: nicolas.le-roux@inserm.fr

Carolina Cabezas

Urs Lucas Böhm

Jean Christophe Poncer

Input-specific learning rules at excitatory synapses onto hippocampal parvalbumin-expressing interneurons

Key points

• Parvalbumin-expressing interneurons represent a major source of inhibition of CA1 hippocampal principal cells and influence both spike timing precision and network oscillations. • These interneurons receive both feed-forward and feedback excitatory inputs which recruit them in the hippocampal network.

• In this study, we compared the functional properties of these two inputs and how they may be modified by neuronal activity.

• We show that calcium-permeable AMPA receptors and NMDA receptors are differentially distributed at feed-forward versus feedback inputs and act as coincidence detectors of opposing modalities.

• Our results reveal that the two major excitatory inputs onto CA1 parvalbumin-expressing interneurons undergo long term plasticity with different frequency regimes of afferent activity, which is likely to influence their function under both normal and pathological conditions.

Introduction

GABAergic inputs onto cortical principal neurons exert both a spatial and temporal control of their activity [START_REF] Isaacson | How inhibition shapes cortical activity[END_REF]. GABAergic interneurons are recruited by excitatory inputs from principal neurons which in turn they inhibit. These excitatory afferents include feed-forward (FF) inputs made by long-range axons from different cortical areas or sub-regions and feedback (FB) inputs from neighbouring principal neurons. Inhibition recruited by FF and FB afferents effectively shapes principal cell activity in a different manner. Disynaptic, FF inhibition enforces precise temporal integration of excitatory inputs onto principal cells by shortening the time window of EPSP summation [START_REF] Pouille | Enforcement of temporal fidelity in pyramidal cells by somatic feed-forward inhibition[END_REF][START_REF] Isaacson | How inhibition shapes cortical activity[END_REF]. In contrast, FB inhibition scales GABAergic inhibition to local excitatory output, thereby contributing to the generation of rhythmic activities [START_REF] Mann | Perisomatic feedback inhibition underlies cholinergically induced fast network oscillations in the rat hippocampus in vitro[END_REF].

In the CA1 area, some GABAergic interneurons participate predominantly in either feed-forward [START_REF] Pouille | Enforcement of temporal fidelity in pyramidal cells by somatic feed-forward inhibition[END_REF] or feedback [START_REF] Maccaferri | Stratum oriens horizontal interneurone diversity and hippocampal network dynamics[END_REF] inhibition. Parvalbumin-expressing interneurons (PV INs), however, fulfil both functions as they receive both FF afferents from Schaffer collaterals and perforant path and FB afferents from neighbouring principal cells [START_REF] Takacs | Extrinsic and local glutamatergic inputs of the rat hippocampal CA1 area differentially innervate pyramidal cells and interneurons[END_REF]. These cells establish predominantly perisomatic contacts onto pyramidal neurons and are therefore well suited for controlling the discharge of their postsynaptic targets [START_REF] Cobb | Synchronization of neuronal activity in hippocampus by individual GABAergic interneurons[END_REF][START_REF] Pouille | Enforcement of temporal fidelity in pyramidal cells by somatic feed-forward inhibition[END_REF]. In addition, their axonal divergence allows them to effectively synchronize large neuronal ensembles [START_REF] Cobb | Synchronization of neuronal activity in hippocampus by individual GABAergic interneurons[END_REF]. The efficacy of excitatory inputs onto PV INs greatly influences their spatial and temporal control over principal cell activity [START_REF] Racz | Augmented hippocampal ripple oscillations in mice with reduced fast excitation onto parvalbumin-positive cells[END_REF][START_REF] Korotkova | NMDA receptor ablation on parvalbumin-positive interneurons impairs hippocampal synchrony, spatial representations, and working memory[END_REF]. Activity-dependent plasticity of synaptic strength at these synapses is then likely to be required to maintain the precisely timed operation of PV INs.

Although long thought to be intrinsically non-plastic [START_REF] Mcbain | Glutamatergic synapses onto hippocampal interneurons: precision timing without lasting plasticity[END_REF], excitatory inputs onto hippocampal interneurons have recently been shown to undergo various forms of long-term, activity-dependent plasticity (Kullmann & Lamsa, 2007;[START_REF] Pelletier | Long-term synaptic plasticity in hippocampal feedback inhibitory networks[END_REF]. Hebbian, NMDAR-dependent LTP has been observed at FF inputs onto some st. radiatum interneurons [START_REF] Lamsa | Hebbian LTP in feed-forward inhibitory interneurons and the temporal fidelity of input discrimination[END_REF] and acts to maintain the temporal fidelity of synaptic integration in pyramidal cells. Another form of LTP depending on activation of calcium-permeable (CP-) AMPARs has been reported in several interneurons subtypes [START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF][START_REF] Oren | Role of ionotropic glutamate receptors in long-term potentiation in rat hippocampal CA1 oriens-lacunosum moleculare interneurons[END_REF][START_REF] Nissen | Cell type-specific long-term plasticity at glutamatergic synapses onto hippocampal interneurons expressing either parvalbumin or CB1 cannabinoid receptor[END_REF][START_REF] Szabo | Calcium-permeable AMPA receptors provide a common mechanism for LTP in glutamatergic synapses of distinct hippocampal interneuron types[END_REF]. Unlike NMDAR-dependent LTP, this form of plasticity requires coincident presynaptic activation and postsynaptic hyperpolarization and was thus named anti-Hebbian [START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]). Thus, opposing learning rules of excitatory inputs onto various interneurons subtypes seem to rely on the relative availability of synaptic NMDA vs. CP-AMPARs (Kullmann & Lamsa, 2007). However, whether this may apply to distinct inputs onto the same interneuron is unknown.

Here, we compared the functional and plastic properties of FF and FB excitatory inputs onto CA1 PV INs. We show these inputs exhibit distinct forms of long term potentiation that reflect a differential distribution of synaptic CP-AMPARs and NMDARs. These two forms of plasticity are induced by different regimes of afferent activity. Thus, distinct postsynaptic receptors at FF vs. FB inputs onto PV interneurons confer these inputs with differential frequency selectivity for long term, activity-dependent adaptation.

Methods

Animals

Pvalb tm1(cre)Arbr mice expressing Cre recombinase under the control of the Pvalb promoter (kindly provided by Dr Silvia Arber, University of Basel, Switzerland; [START_REF] Hippenmeyer | A developmental switch in the response of DRG neurons to ETS transcription factor signaling[END_REF] were crossed with RCE:LoxP reporter mice expressing enhanced green fluorescent protein (EGFP) under the control of promoter sequences for the Rosa locus enhanced with an additional CAG sequence (provided by Dr Gord Fishell, NYU, USA; [START_REF] Sousa | Characterization of Nkx6-2-derived neocortical interneuron lineages[END_REF]. The genetic background of both Pvalb tm1(cre)Arbr and RCE:LoxP mice was C57Bl6(J). Homozygous, PV Cre ::RCE mice were used in all experiments. All the procedures carried out conformed to the International Guidelines on the ethical use of animals.

C 2013 The Authors. The Journal of Physiology C 2013 The Physiological Society

Electrophysiology

Horizontal hippocampal slices (300-350 μm thick) were prepared from postnatal day 17-23 PV Cre ::RCE mice as described [START_REF] Scheuber | Presynaptic Cav2.1 and Cav2.2 differentially influence release dynamics at hippocampal excitatory synapses[END_REF]. Mice were anesthetized by intraperitoneal injection of ketamine and xylazine (80 and 20 mg kg -1 , respectively) and killed by decapitation. The brain was quickly removed and immersed in low-sodium, ice-cold artificial CSF (low Na + -ACSF) equilibrated with 95% O 2 -5% CO 2 . The composition of the low Na + -ACSF was as follows: (in mM) 248 sucrose, 26 NaHCO 3 , 10 glucose, 5 MgCl 2 , 4 KCl, 1 CaCl 2 , and 0.005 Phenol Red. After complete surgical (from st. lacunosum down to the alveus) transection between CA3 and CA1 areas to prevent propagation of recurrent excitation along Schaffer collaterals, slices were transferred to a submerged chamber maintained at 31 • C, mounted on an upright microscope (BX51WI, Olympus France SA, Rungis, France) and superfused at 2.5 ml min -1 with ACSF composed of (in mM): 124 NaCl, 26.2 NaHCO 3 , 11 D-glucose, 2.5 KCl, 1 NaH 2 PO 4 , 2 CaCl 2 and 3 MgCl 2 .

Whole-cell patch-clamp recordings were made using borosilicate glass microelectrodes (3-5 M resistance) filled with (in mM): 105 CsMeSO 3 , 20 CsCl, 10 Hepes, 10 EGTA, 4 MgATP, 0.4 Na 3 GTP (pH 7.4, 291 mosmol l -1 ). For long term plasticity experiments, EGTA concentration was reduced to 0.1 mM and spermine was added to the internal solution to better preserve the voltage-dependent block of CP-AMPARs [START_REF] Bowie | Inward rectification of both AMPA and kainate subtype glutamate receptors generated by polyamine-mediated ion channel block[END_REF][START_REF] Donevan | Intracellular polyamines mediate inward rectification of Ca(2+)-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors[END_REF][START_REF] Koh | Block of native Ca 2+ -permeable AMPA receptors in rat brain by intracellular polyamines generates double rectification[END_REF]. EPSCs were evoked at 0.1 Hz in the presence of bicuculline methochloride (20 μM) using ortho-or antidromic stimulation through monopolar glass microelectrodes placed either in st. radiatum to recruit feed-forward (FF) Schaffer collateral inputs and/or in st. oriens/alveus to recruit predominantly feedback (FB) inputs. Stimulation intensity and duration typically ranged 50-250 μA and 100-150 μs, respectively, for both interneuron and pyramidal cell recordings.

In most experiments (Fig. 2345), synaptic plasticity was induced in voltage clamp by pairing 400 presynaptic stimuli delivered at 5 Hz with either continuous hyperpolarization to -90 mV (for anti-Hebbian conditioning) or depolarization to 0 mV (for Hebbian conditioning) [START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]. For all recordings, pairing was applied less than 10 min after break in (typically 5), in order to prevent washout of the intracellular milieu [START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]. In some experiments (Fig. 6), we tested the plasticity induced by various frequency regimes of afferent activity (900 pulses at 0.1, 1, 5 or 20 Hz [START_REF] Dudek | Homosynaptic long-term depression in area CA1 of hippocampus and effects of N-methyl-D-aspartate receptor blockade[END_REF] or 100 pulses at 100 Hz repeated 5 times 10 s apart [START_REF] Lamsa | Hebbian LTP in feed-forward inhibitory interneurons and the temporal fidelity of input discrimination[END_REF][START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]). In these experiments, EPSCs were recorded in voltage-clamp mode, but the conditioning protocol was delivered in current clamp from resting membrane potential (-58.8 ± 0.4 mV for PV INs, -58.5 ± 1.3 mV for pyramidal cells). Neurons were therefore recorded with a potassium-based internal solution containing (in mM): 120 KMeSO 3 , 10 KCl, 10 Hepes, 0.1 EGTA, 4 MgATP, 0.4 Na3GTP and 0.1 spermine (pH 7.4, 292 mosmol l -1 ).

Glutamate uncaging onto individual PV+ INs was performed in the presence of 200-300 μM MNI-glutamate (Tocris Bioscience, Bristol, UK) together with bicuculline and tetrodotoxin using a 405 nm, laser diode beam (Deepstar, Omicron, Photon Lines, Saint-Germain-en-Laye, France) conducted through a multimode optic fibre and alignment device (Prairie Technologies, Middleton, WI, USA) set to generate a <3 μm spot in the objective focus. The power of the laser head output was controlled using Omicron Laser Controller v2.97, while trigger and pulse duration were set using Clampex and a Digidata controller. The stimulation site was set along the primary dendrite of a PV IN to obtain the fastest uEPSC onset kinetics with minimal laser intensity (2-20 mW) in order to minimize recruitment of extrasynaptic receptors. Pre-induction recordings were performed at 0.1 Hz with stimulation duration of 1.5 ms for at least 5 min. A pairing protocol was then applied, consisting of 100 pulses at 5 Hz while holding the cell at -90 mV. During the induction the stimulation duration was increased to 4 ms. uEPSCs were then recorded in the same conditions as prior to pairing for at least another 30 min.

Firing properties of PV INs were recorded in current-clamp mode using an internal solution containing (in mM): 120 KMeSO 3 , 10 KCl, 10 Hepes, 0.1 EGTA, 4 MgATP and 0.4 Na 3 GTP (pH 7.4, 292 mosmol l -1 ). To generate firing frequency = f(current) curves, a range of depolarizing current pulses (0 to 0.6 nA) were delivered with an interstimulus interval of 2 s.

Signals were acquired and filtered at 10 kHz using a Multiclamp amplifier and digitized at 20 kHz using Clampex software (Molecular Devices, St. Grégoire, France). All parameters were analysed offline with Clampfit. Peak EPSC amplitudes were always measured between 1.8 and 2.8 ms of stimulation artefact to prevent contamination by disynaptic responses. Bicuculline, D,L-APV, Ro25-6981, DPCPX, NBQX were all purchased from Ascent Scientific (abcamBiochemicals, Cambridge, UK). Naspm was obtained from Tocris Bioscience.

Immunohistochemistry

P21 PV Cre ::RCE mice were rapidly anaesthetized by intraperitoneal injection of ketamine and xylazine (80 and 20 mg kg -1 , Sigma-Aldrich) and perfused transcardially with 4% (w/v) paraformaldehyde in 0.1 M sodium phosphate buffer, pH 7.5. Brains were post-fixed overnight in the same solution, stored at 4 • C and cryoprotected J Physiol 00.00 in 30% sucrose for an additional 48 h. Transverse, 40 μm-thick sections were cut with a cryotome (Microm KS34, Thermo Scientific Saint Herblain, France) and stored at -20 • C in a solution containing 30% (v/v) ethylene glycol, 30% (v/v) glycerol, and 0.1 M sodium phosphate buffer, until they were processed for immunofluorescence. Free-floating sections were rinsed in PBS and then incubated for 1 h in PBS supplemented with 0.3% Triton X-100 and 5% normal goat serum. They were then incubated overnight at 4 • C with primary antibodies. The following antibodies were used: GFP (polyclonal chicken, 1:1000, Chemicon, Millipore France SAS, Saint Quentin en Yvelines, France); PV (polyclonal rabbit, 1:1000, Swant, Marly, Switzerland). Sections were then rinsed in PBS and incubated for 1 h with donkey-FITC (1:500; all from Jackson Laboratory, Bar Harbor, ME, USA) and goat Cy3-coupled (1:600) secondary antibodies. Sections were rinsed in PBS and mounted with Mowiol/Dabco (25 mg ml -1 ) and stored at 4 • C before imaging.

Acquisition and analysis of images

Double-labelled images were obtained using fluorescence microscopy (DM6000; Leica Microsystems SAS, Nanterre, France) using × 10 and × 40 objectives. For estimating the proportion of GFP+ and PV+ cells, the entire CA1 was imaged in, at least, three different slices from each animal. The GFP+ and PV+ cells were counted manually.

Data are presented as means ± SEM. Statistical significance was assessed using a Mann-Whitney or Wilcoxon tests using SigmaStat (RITME, Paris, France). Principal component analysis (see online Supplementary material Fig. S1) was performed using Xlstat software (Addinsoft, Paris, France) under Microsoft Excel.

Results

In order to facilitate recordings from PV INs, we generated a mouse strain expressing EGFP specifically in these cells, by crossing Pvalb tm1(cre)Arbr mice [START_REF] Hippenmeyer | A developmental switch in the response of DRG neurons to ETS transcription factor signaling[END_REF] with the sensitive reporter strain RCE:LoxP [START_REF] Sousa | Characterization of Nkx6-2-derived neocortical interneuron lineages[END_REF]. As expected from the delayed postnatal expression of PV in cortical interneurons [START_REF] Solbach | Ontogeny of the calcium binding protein parvalbumin in the rat nervous system[END_REF], EGFP was detected in hippocampal interneurons from homozygous Pvalb tm1(cre)Arbr ::RCE:LoxP mice (referred to as PV Cre ::RCE thereafter) from stages P7-P10. In area CA1, EGFP expression was detected in 93.0 ± 1.5% of PV-immunopositive neurons and 97.5 ± 0.3% EGFP-immunopositive interneurons were also immunopositive for PV (Supplementary Fig. S1A andB). Most (8 out of 11) neurons displayed fast spiking properties, short-lasting action potentials and fast AHP typical of hippocampal PV INs (Supplementary Fig. S1C andD). In addition, we could not distinguish sub-groups of PV INs based on their intrinsic physiological properties (Supplementary Fig. S1E andF). Therefore, EGFP expression faithfully reflects PV expression and PV Cre ::RCE mice represent a valuable model for studying the physiology of hippocampal PV INs.

Distinct postsynaptic receptors at feed-forward vs. feedback excitatory inputs onto PV INs

Long term synaptic plasticity in hippocampal interneurons relies primarily on activation of Ca 2+ -permeable postsynaptic glutamate receptors including GluA2-lacking AMPA receptors and NMDA receptors (Kullmann & Lamsa, 2007). Therefore we first compared the properties of EPSCs at feed-forward (FF) and feedback (FB) inputs onto CA1 PV INs. Whole-cell patch-clamp recordings were made from EGFP expressing cells with their soma within st. pyramidale in the CA1 area (Fig. 1A). EPSCs were evoked by focal stimulation either in st. radiatum to recruit FF inputs from Schaffer collaterals or in the st. oriens/alveus to recruit predominantly FB inputs from neighbouring CA1 pyramidal neurons [START_REF] Takacs | Extrinsic and local glutamatergic inputs of the rat hippocampal CA1 area differentially innervate pyramidal cells and interneurons[END_REF]; Fig. 1B). Pharmacologically isolated AMPAR-mediated EPSCs evoked from either pathway onto PV INs showed pronounced, although not complete, inward rectification (Fig. 1C andD), suggesting they were carried predominantly by receptors lacking the GluA2 subunit. Consistent with this conclusion, AMPAR-mediated EPSCs in PV INs were partially blocked by the selective antagonist of Ca 2+ -permeable (CP-) AMPARs, 1-Naphthyl acetyl spermine trihydrochloride (Naspm; [START_REF] Koike | Blocking effect of 1-naphthyl acetyl spermine on Ca(2+)-permeable AMPA receptors in cultured rat hippocampal neurons[END_REF]. Naspm (100 μM) reduced EPSC amplitude to a similar extent at FF and FB inputs onto PV INs (to 36.3 ± 3.3 and 40.0 ± 3.9% of control, respectively, P = 0.6, n = 10) but not at FF inputs onto principal cells (109.1 ± 5.2% of control, n = 2; Fig. 1E andF).

In hippocampal interneurons, synapses expressing CP-AMPARs classically display small NMDAR-mediated EPSC components [START_REF] Lei | Distinct NMDA receptors provide differential modes of transmission at mossy fiber-interneuron synapses[END_REF]Kullmann & Lamsa, 2007;[START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]). Quantitative immunogold studies have suggested that excitatory synapses onto hippocampal PV INs may even be largely devoid of NMDARs [START_REF] Nyiri | Large variability in synaptic N-methyl-D-aspartate receptor density on interneurons and a comparison with pyramidal-cell spines in the rat hippocampus[END_REF]. Consistent with this observation, the NMDA/AMPA ratio of EPSCs evoked by Schaffer collateral stimulation in PV INs was considerably smaller than in neighbouring pyramidal cells (0.16 ± 0.02 vs. 1.23 ± 0.01; P < 0.001; Fig. 1G). However, this ratio was significantly greater for EPSCs evoked at FB compared to FF inputs onto PV INs (0.28 ± 0.04 vs. 0.16 ± 0.02, P < 0.05, n = 12 cells) but not pyramidal cells (1.11 ± 0.24 vs. 1.23 ± 0.01, P = 0.6, n = 8 cells). The amplitude of the NMDAR-mediated component of EPSCs evoked at distinct inputs onto CA3 interneurons has been correlated with their content in GluN2A and GluN2B subunits [START_REF] Lei | Distinct NMDA receptors provide differential modes of transmission at mossy fiber-interneuron synapses[END_REF]. We therefore asked whether the differential -37 ± 10% of control, P = 0.9, n = 9 cells; Fig. 1H). This effect, however, was more modest at excitatory synapses onto pyramidal cells (-17 ± 4% of control, P = 0.07, n = 12 cells). Thus, activation of both FB and FF excitatory synapses onto PV INs recruits GluA2-lacking AMPARs but NMDARs are more abundant at FB than at FF synapses.

Plasticity rules at FF vs. FB excitatory inputs onto PV interneurons

CP-AMPARs and NMDARs both act as coincidence detectors yet of opposing modalities. Ca 2+ influx through the former is maximal during coincident glutamate release and postsynaptic hyperpolarization which both prevents channel blockade by endogenous polyamines and increases the driving force for Ca 2+ ions. In contrast, Ca 2+ influx through NMDARs requires coincident glutamate release and postsynaptic depolarization to relieve channel blockade by Mg 2+ ions. We therefore asked whether the differential endowment of FF and FB inputs onto PV INs with postsynaptic receptors may influence plasticity rules at these synapses. We recorded EPSCs evoked by either FF or FB stimulation (at 0.1 Hz) before and after low-frequency pairing (400 pulses at 5 Hz) of only one pathway (Fig. 2). Two pairing modalities were tested: a Hebbian protocol consisting in pairing afferent stimulation with postsynaptic depolarization to 0 mV, and an anti-Hebbian protocol where presynaptic activation was paired with postsynaptic hyperpolarization to -90 mV [START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]). Anti-Hebbian pairing induced a persistent potentiation of both FF and FB inputs (+45.5 ± 14.5 and +38.8 ± 19.9% of control EPSC amplitude, respectively; n = 11 and 9 cells; Fig. 2A andB), which lasted for as long as recordings were maintained (>40 min). This form of potentiation was input specific as the other, unpaired pathway showed no significant plasticity (-2.6 ± 2.2 and +5.6 ± 1.2% on unpaired FB and FF inputs, respectively). In contrast, Hebbian pairing produced no long-lasting plasticity of FF inputs (+3.6 ± 6.2% of control EPSC amplitude) but a persistent potentiation of FB inputs (+38.4 ± 18.4%, n = 8; Fig. 2C andD). Thus, FF and FB excitatory inputs onto PV INs express two forms of LTP with opposing induction modalities which correlate with the differential expression of postsynaptic CP-AMPARs and NMDARs.

We therefore tested whether anti-Hebbian and Hebbian LTP induction in PV INs indeed relied on activation of CP-AMPARs and NMDARs, respectively. We used Naspm (100 μM) to partially block EPSCs immediately prior to delivering an anti-Hebbian pairing protocol and compared the recovery of the EPSC upon washing out Naspm in control vs. paired pathways (Fig. 3A). Naspm completely prevented anti-Hebbian LTP both at FF and FB inputs onto PV INs (Fig. 3D; n = 6 cells each). In contrast, application of APV (100 μM) did not block this form of plasticity (Fig. 3C; n = 10 and 11 cells, respectively) but abolished Hebbian LTP at FB inputs onto PV INs (Fig. 3B andD; n = 14 cells). These results demonstrate that in PV INs, two opposing forms of LTP are induced by activation of distinct postsynaptic receptor subtypes: anti-Hebbian LTP is mediated primarily by CP-AMPARs whereas Hebbian LTP relies on activation of NMDARs.

LTP expression in PV interneurons

We next examined the locus of LTP expression in PV INs using three independent approaches. We first tested whether LTP in PV INs was associated with changes in the short term plasticity of excitatory inputs. As previously reported in some unidentified basket or perisomatic CA2 and CA1 interneurons [START_REF] Losonczy | Cell type dependence and variability in the short-term plasticity of EPSCs in identified mouse hippocampal interneurones[END_REF][START_REF] Pouille | Routing of spike series by dynamic circuits in the hippocampus[END_REF], paired stimuli delivered 50 ms apart evoked EPSCs with a mean paired-pulse ratio (PPR) of 0.90 ± 0.05 at both FF and FB synapses (n = 15 and 24 cells, respectively), comparable to that described at unitary connections [START_REF] Ali | Facilitating pyramid to horizontal oriens-alveus interneurone inputs: dual intracellular recordings in slices of rat hippocampus[END_REF][START_REF] Mercer | Local circuitry involving parvalbumin-positive basket cells in the CA2 region of the hippocampus[END_REF]. The PPR of evoked EPSCs was not significantly changed upon anti-Hebbian LTP at FF (-4.9 ± 4.1%, P = 0.6, n = 15 cells) or FB synapses (-5.0 ± 2.9%, P = 0.6, n = 14 cells) or upon Hebbian LTP at FB synapses onto PV INs (-2.5 ± 3.0%, P = 0.7, n = 10 cells; Fig. 4A andB). These results suggest both forms of LTP may not primarily reflect changes in transmitter release. We next monitored the rate of EPSC suppression by the CP-AMPAR blocker Naspm as an index of release probability [START_REF] Mainen | Use-dependent AMPA receptor block in mice lacking GluR2 suggests postsynaptic site for LTP expression[END_REF]. Inhibition of presynaptic A1 receptor is known to potentiate glutamatergic inputs onto hippocampal interneurons [START_REF] Doherty | Regulation of excitatory input to inhibitory interneurons of the dentate gyrus during hypoxia[END_REF]. Application of the selective A1R antagonist DCPCX (100 μM) indeed potentiated EPSCs evoked at FF inputs onto PV INs by 43.3 ± 11.6% (P < 0.01, n = 10 cells; Fig. 4C andD). Subsequent application of Naspm produced a use-dependent block of EPSCs with a decay time constant of 3.5 ± 0.6 s (n = 9 cells), significantly faster than in interleaved, control experiments in the absence of DPCPX (5.5 ± 0.7 s, P < 0.05, n = 17 cells). Thus, a ∼40% potentiation of purely presynaptic origin at excitatory inputs on PV INs can be readily detected as an increase in the rate of Naspm-induced blockade of EPSCs. We then repeated the same experiment while comparing two independent FF inputs onto PV INs. Anti-Hebbian pairing of one pathway produced a 39.6 ± 8.5% increase in EPSC amplitude with no significant change in the other, control pathway (-4.4 ± 5.6%, n = 21 cells). Application of Naspm 15 min after pairing induced a blockade of EPSCs at a rate that was not significantly different between the two pathways (P = 0.8, n = 21 cells; Fig. 4E andF).

Noticeably, the proportion of EPSC blocked by Naspm was also not significantly different between the potentiated and the control pathways (0.38 ± 0.05 vs. 0.43 ± 0.06 of control amplitude, respectively, P = 0.5). Together, these results strongly argue against LTP of excitatory inputs onto PV INs being expressed as a persistent change in release probability.

We therefore asked whether LTP of excitatory synapses onto PV INs could undergo long term plasticity independent of presynaptic function. We used laser-based, single-photon photolysis of MNI-glutamate (see Methods) to induce uncaging-evoked excitatory postsynaptic currents (uEPSCs) in EGFP+ PV INs. The position of the laser spot (≈ 3 μm in diameter) was adjusted along proximal dendrites in st. radiatum (20-40 μm from soma) to induce uEPSCs at 0.1 Hz that were fast rising and comparable in amplitude to mEPSCs (22.7 ± 2.2 vs. 18.3 ± 1.6 pA, n = 11 and 19 cells, respectively, P = 0.1; Fig. 5A). The peak amplitude and kinetics of uEPSCs were strongly dependent on subtle variations in the position of the laser (Fig. 5B), suggesting these currents might reflect activation of synaptic sites. Anti-Hebbian pairing (100 stimuli delivered at 5 Hz while holding the recorded cell at -90 mV) was then applied within less than 10 min after break-in and led to a persistent (>30 min) potentiation of uEPSC amplitude ranging from 25 to 255% (average, +83.2 ± 27.9% of control, n = 8 cells; Fig. 5C andD). Meanwhile, uEPSCs evoked at unpaired loci remained unaffected (-4.3 ± 3.6% of control amplitude, n = 3 cells). Similarly, Hebbian pairing (same train of stimuli delivered while holding the cell at 0 mV) led to a persistent potentiation of uEPSC evoked by glutamate uncaging onto distal (>50 μm from soma, close to alveus border) but not proximal (20-40 μm from soma) dendrites in st. oriens (+73.3 ± 19.9% of control, n = 7 cells, and +3.5 ± 10.1% of control, n = 6 cells respectively, Fig. 5E andF). We conclude that excitatory synapses onto PV INs can undergo homosynaptic LTP that relies exclusively on postbut not presynaptic modifications.

Frequency dependence of long term plasticity at FF vs. FB inputs onto PV INs

Our results show that synaptic activation of CP-AMPARs and NMDARs underlie two forms of long term plasticity that are differentially induced at FF and FB inputs onto PV INs. Under which regimes of afferent activity may synaptic efficacy at these inputs be selectively modified? During repetitive afferent activity, EPSP integration leads to a progressive depolarization of the postsynaptic membrane. This depolarization may lead to a frequency-dependent blockade of CP-AMPARs by endogenous polyamines whereas NMDAR activation may be facilitated due to relief of Mg 2+ block. Thus, the sign and amplitude of CP-AMPAR-vs. NMDAR-dependent plasticity may be differentially influenced by the frequency of afferent stimulation [START_REF] Dudek | Homosynaptic long-term depression in area CA1 of hippocampus and effects of N-methyl-D-aspartate receptor blockade[END_REF]. We tested this hypothesis by comparing synaptic plasticity of FF and FB afferents onto PV INs induced by trains of afferent stimuli delivered at varying frequencies. Cells were held around their resting membrane potential in current-clamp mode and 1-100 Hz afferent stimulation was delivered to either FF or FB inputs (see Methods; Fig. 6). At frequencies up to 5 Hz, no summation of EPSPs was detected (Fig. 6A andB). At 100 Hz, however, summated EPSPs often depolarized PV INs above threshold, leading to the firing of action potentials (Fig. 6D andE). Unlike in pyramidal cells, low frequency stimulation did not lead to detectable LTD of excitatory inputs onto PV INs. However, 20 Hz stimulation induced a more pronounced LTP at synapses onto PV INs than that onto pyramidal cells (P < 0.05; Fig. 6C andG). Most strikingly, 100 Hz afferent stimulation led to a robust LTP of EPSCs both at Schaffer collateral synapses onto pyramidal cells (+123 ± 33% of control, n = 5 cells) and FB inputs onto PV INs (+144 ± 42% of control, n = 5 cells) whereas no potentiation was induced at FF inputs onto PV INs (+5.3 ± 17% of control, n = 7 cells; Fig. 6D andG). We asked whether these distinct behaviours of FF and FB inputs upon 100 Hz stimulation reflected a differential recruitment of NMDA vs. CP-AMPARs. Application of 100 μM APV almost completely abolished the potentiation of EPSCs upon 100 Hz stimulation of FB afferents onto PV INs (to +13.8 ± 9.0% of control, n = 7 cells; Fig. 6E andG), suggesting LTP at this frequency primarily relied on NMDAR activation. Conversely, 100 Hz stimulation of FF inputs induced a robust potentiation (to +87.4 ± 23.1% of control, n = 6 cells; Fig. 6F andG) when delivered while holding PV INs at -90 mV to maximize Ca 2+ influx through CP-AMPARs. Thus, our results show that the distinct endowment of excitatory synapses onto PV INs with various, calcium-permeable glutamate receptors accounts for a differential, long-term adaptation to specific frequency regimes of afferent activity.

Discussion

We have shown that distinct excitatory inputs onto CA1 PV INs exhibit different rules of long term plasticity depending on distinct postsynaptic receptors. The specific conditions required for CP-AMPAR and N. Le Roux and others J Physiol 00.00 NMDAR activation endow these inputs with a differential frequency selectivity of long term plasticity induction.

PV-expressing INs comprise several neuron subtypes targeting distinct membrane compartments on their postsynaptic targets. In particular, PV expression is found in basket and axo-axonic cells as well as in bistratified cells and, to a lesser extent, oriens-lacunosum moleculare (O-LM) cells [START_REF] Klausberger | Neuronal diversity and temporal dynamics: the unity of hippocampal circuit operations[END_REF]. Since all recordings in this study were performed from PV INs with their soma within or close to st. pyramidale, it seems unlikely that O-LM cells were recorded. Although basket cells represent the majority of PV INs, we cannot exclude, however, the possibility that other PV INs were included in this study. In fact, three out of eleven PV INs did not show typical fast-spiking properties. Yet, no significant variability in the synaptic or plastic properties of PV INs was detected in this study. Thus, excitatory inputs onto various PV INs may share common plasticity rules or we may have oversampled one specific subtype of PV INs in our experiments. In the CA1 area, PV INs receive both FF and FB excitatory inputs. FF inputs include Schaffer collaterals from CA3 principal neurons as well as entorhinal inputs from the perforant path and inputs from the thalamus and the amygdala, whereas FB inputs originate from neighbouring pyramidal neurons [START_REF] Klausberger | Neuronal diversity and temporal dynamics: the unity of hippocampal circuit operations[END_REF]. These inputs are differentially segregated among the hippocampal layers such that most excitatory inputs in st. radiatum originate from Schaffer collaterals. Glutamatergic synapses in st. oriens/alveus are however more heterogeneous. Nevertheless, more than 50% recurrent CA1 excitatory inputs and less than 20% direct entorhinal inputs in this layer innervate local interneurons [START_REF] Takacs | Extrinsic and local glutamatergic inputs of the rat hippocampal CA1 area differentially innervate pyramidal cells and interneurons[END_REF]. Therefore, although we cannot exclude that stimulation in st. oriens/alveus may have recruited glutamatergic synapses of various origins, it seems reasonable to assume that most of them originate from recurrent CA1 axons [START_REF] Pouille | Routing of spike series by dynamic circuits in the hippocampus[END_REF]. Thus, EPSCs evoked from st. radiatum and st. oriens/alveus in PV INs predominantly originate from FF and FB afferents, respectively.

Our results show that FF and FB inputs onto PV INs express different sets of postsynaptic glutamate receptors. AMPARs at excitatory inputs onto several hippocampal interneurons including PV INs have been described as predominantly lacking the GluA2 subunit [START_REF] Leranth | AMPA receptors in the rat and primate hippocampus: a possible absence of GluR2/3 subunits in most interneurons[END_REF][START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF][START_REF] Nissen | Cell type-specific long-term plasticity at glutamatergic synapses onto hippocampal interneurons expressing either parvalbumin or CB1 cannabinoid receptor[END_REF]. Consistent with these reports, we observed strong inward rectification of AMPAR-mediated EPSCs and partial block by the specific, GluA2-lacking AMPAR blocker Naspm. These properties were similar at FF and FB inputs onto PV INs and are likely to reflect a partial voltage-dependent blockade by endogenous cytoplasmic polyamines [START_REF] Bowie | Inward rectification of both AMPA and kainate subtype glutamate receptors generated by polyamine-mediated ion channel block[END_REF][START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]. NMDARs are instead differentially expressed at synapses from FF vs. FB excitatory inputs onto PV INs. EPSCs evoked upon stimulation of FF inputs are largely devoid of an NMDAR-mediated component. In contrast, EPSCs evoked at FB inputs show a significantly larger NMDA/AMPA ratio, although not as large as that of EPSCs recorded in principal neurons. Studies based on immunogold staining of the GluN1 subunit in rat hippocampus have revealed an almost complete absence of GluN1 at asymmetrical synapses onto PV IN dendrites [START_REF] Nyiri | Large variability in synaptic N-methyl-D-aspartate receptor density on interneurons and a comparison with pyramidal-cell spines in the rat hippocampus[END_REF]. Interestingly, however, this observation was made on dendritic branches within st. radiatum. Instead, synapses established onto dendrites from unidentified interneurons within st. oriens showed an intermediate density of gold particles per synapses as compared to pyramidal cell dendrites and st. radiatum dendrites of PV INs [START_REF] Nyiri | Large variability in synaptic N-methyl-D-aspartate receptor density on interneurons and a comparison with pyramidal-cell spines in the rat hippocampus[END_REF]. Thus, immunogold data and our present results converge to suggest a differential expression of GluN1 at FF vs. FB inputs onto PV INs. A similar, input-specific distribution of postsynaptic glutamate receptors has been observed in CA3 interneurons [START_REF] Toth | Afferent-specific innervation of two distinct AMPA receptor subtypes on single hippocampal interneurons[END_REF][START_REF] Lei | Distinct NMDA receptors provide differential modes of transmission at mossy fiber-interneuron synapses[END_REF]. However, in contrast to these cells, the synaptic CP-AMPAR content at synapses onto CA1 PV INs does not appear correlated with NMDAR content. The mechanisms that govern the selective targeting of NMDARs to FB inputs onto these cells now remain to be elucidated.

We report that AMPARs and NMDARs confer FF and FB inputs onto PV INs with distinct forms of long term plasticity. Both AMPAR-and NMDAR-mediated LTP have been reported at excitatory synapses onto CA1 hippocampal interneurons (Kullmann & Lamsa, 2007). However, either form of plasticity was usually observed in specific interneuron sub-populations. NMDAR-mediated LTP has been described primarily in st. radiatum interneurons mediating FF inhibition [START_REF] Lamsa | Hebbian LTP in feed-forward inhibitory interneurons and the temporal fidelity of input discrimination[END_REF] whereas AMPAR-mediated LTP was observed in several interneuron subtypes [START_REF] Oren | Role of ionotropic glutamate receptors in long-term potentiation in rat hippocampal CA1 oriens-lacunosum moleculare interneurons[END_REF][START_REF] Nissen | Cell type-specific long-term plasticity at glutamatergic synapses onto hippocampal interneurons expressing either parvalbumin or CB1 cannabinoid receptor[END_REF][START_REF] Szabo | Calcium-permeable AMPA receptors provide a common mechanism for LTP in glutamatergic synapses of distinct hippocampal interneuron types[END_REF] including at FB inputs onto CA1 PV INs [START_REF] Nissen | Cell type-specific long-term plasticity at glutamatergic synapses onto hippocampal interneurons expressing either parvalbumin or CB1 cannabinoid receptor[END_REF]). In the latter experiments, however, NMDARs were pharmacologically blocked, thereby probably masking their contribution to Hebbian plasticity. Group I mGluRs have also been involved in the induction of both Hebbian [START_REF] Perez | A hebbian form of long-term potentiation dependent on mGluR1a in hippocampal inhibitory interneurons[END_REF]) and anti-Hebbian (Le [START_REF] Duigou | Group I mGluR agonist-evoked long-term potentiation in hippocampal oriens interneurons[END_REF] forms of LTP in CA1 interneurons. Since PV INs express mGluR5 [START_REF] Van Hooft | Differential expression of group I metabotropic glutamate receptors in functionally distinct hippocampal interneurons[END_REF], we cannot exclude the possibility that these receptors may influence the forms of LTP we report. However, our experiments using Naspm demonstrate that specific activation of CP-AMPARs is strictly required for LTP induction using an anti-Hebbian protocol. Conversely, LTP induction with a Hebbian protocol strictly requires NMDAR activation.

CP-AMPARs and NMDARs are classically viewed as coincidence detectors of opposing modalities. Thus, membrane depolarization is required to relieve NMDAR channel blockade by Mg 2+ ions [START_REF] Nowak | Magnesium gates glutamate-activated channels in mouse central neurones[END_REF] whereas blockade of CP-AMPAR channels by endogenous polyamines is relieved by membrane hyperpolarization which also enhances the driving force for Ca 2+ [START_REF] Bowie | Inward rectification of both AMPA and kainate subtype glutamate receptors generated by polyamine-mediated ion channel block[END_REF][START_REF] Donevan | Intracellular polyamines mediate inward rectification of Ca(2+)-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors[END_REF]. A protocol associating afferent stimulation and membrane hyperpolarization may then promote CP-AMPAR-mediated plasticity induction. This type of paradigm was named anti-Hebbian [START_REF] Lamsa | Anti-Hebbian long-term potentiation in the hippocampal feedback inhibitory circuit[END_REF]) as opposed to the classical Hebbian rule for plasticity induction. However, our experiments where LTP was induced in current-clamp mode reveal that membrane hyperpolarization is not required for induction of CP-AMPAR-mediated LTP. In fact, plasticity was readily induced around resting membrane potential up to 20 Hz afferent stimulation which led to significant depolarization due to EPSP summation. Thus, induction of long term plasticity mediated by CP-AMPARs does not strictly require membrane hyperpolarization or depolarization and may thus be called non-Hebbian rather than anti-Hebbian. Nevertheless, membrane potential influences plasticity rules at synapses expressing CP-AMPARs, in particular the frequency range of afferent activity that triggers plasticity (Fig. 6) and possibly the modification threshold as defined in the Bienenstock-Cooper-Munro formalism [START_REF] Bienenstock | Theory for the development of neuron selectivity: orientation specificity and binocular interaction in visual cortex[END_REF][START_REF] Bear | A physiological basis for a theory of synapse modification[END_REF].

Although the mechanisms of LTP in principal cells have been studied in exquisite detail (Malinow & Malenka, J Physiol 00.00 2002;[START_REF] Poncer | Hippocampal long term potentiation: silent synapses and beyond[END_REF], LTP expression in hippocampal interneurons remains poorly understood. In O-LM interneurons, anti-Hebbian LTP has been suggested to be expressed presynaptically, based on concomitant changes in the coefficient of variation (CV) of the EPSPs [START_REF] Oren | Role of ionotropic glutamate receptors in long-term potentiation in rat hippocampal CA1 oriens-lacunosum moleculare interneurons[END_REF]. mGluR-dependent LTP in unidentified st. oriens interneurons on the other hand is associated with both reduced failure rate [START_REF] Lapointe | Synapse-specific mGluR1-dependent long-term potentiation in interneurones regulates mouse hippocampal inhibition[END_REF]) and increased synaptic potency [START_REF] Perez | A hebbian form of long-term potentiation dependent on mGluR1a in hippocampal inhibitory interneurons[END_REF], suggestive of concomitant pre-and postsynaptic modifications. However, changes in both CV and failure rate may be erroneously attributed to presynaptic modifications [START_REF] Poncer | Postsynaptic conversion of silent synapses during LTP affects synaptic gain and transmission dynamics[END_REF]. Our data argue against a presynaptic locus of LTP expression in CA1 PV INs. First, CP-AMPAR and NMDAR-mediated forms of LTP were not associated with a consistent increase in release probability, as detected either by changes in paired-pulse ratio or rate of Naspm-induced blockade of CP-AMPARs. Second, persistent LTP of uEPSCs could be induced by pairing glutamate uncaging onto st. radiatum or st. oriens/alveus dendrites of PV INs with either membrane hyperpolarization or depolarization, respectively (Fig. 5), suggesting both anti-Hebbian and Hebbian LTP may be induced through purely postsynaptic mechanisms in PV INs. Hence we suggest LTP in CA1 PV INs may be primarily expressed postsynaptically, possibly through an increase in postsynaptic receptor number or unitary conductance, as described in principal cells [START_REF] Poncer | Hippocampal long term potentiation: silent synapses and beyond[END_REF]. Although the mechanisms of such postsynaptic LTP expression remain to be elucidated, it is noteworthy that the fraction of AMPAR-mediated EPSC blocked by Naspm was unchanged after anti-Hebbian pairing (Fig. 4E). Therefore expression of this form of LTP at least may not involve changes in subunit composition of postsynaptic AMPARs [START_REF] Plant | Transient incorporation of native GluR2-lacking AMPA receptors during hippocampal long-term potentiation[END_REF].

Our data show that FF and FB excitatory inputs onto PV INs show distinct learning rules that rely on activation of different sets of postsynaptic receptors. These rules confer on FF and FB inputs a specific dependence on afferent activity frequency. Thus, at resting membrane potential, FF inputs are potentiated within a narrower frequency range of afferent activity frequency (5-20 Hz in our experimental conditions) than FB inputs (up to 100 Hz). How may these distinct learning rules influence network operation in the CA1 area? Even moderate changes in the excitatory drive of PV INs are sufficient to alter spike timing precision of PV INs with respect to gamma rhythm, reduce the power of gamma oscillations [START_REF] Fuchs | Recruitment of parvalbumin-positive interneurons determines hippocampal function and associated behavior[END_REF]) and increase burst firing in principal cells [START_REF] Racz | Augmented hippocampal ripple oscillations in mice with reduced fast excitation onto parvalbumin-positive cells[END_REF]. Therefore alterations of synaptic potency at excitatory inputs onto PV INs greatly influence their contribution to the generation of coordinated ensemble activities in the CA1 region. The net impact of the plasticity processes we describe here is likely to depend on PV IN subtypes, which comprise a majority of basket and chandelier cells as well as some bistratified cells [START_REF] Klausberger | Neuronal diversity and temporal dynamics: the unity of hippocampal circuit operations[END_REF]. However, if we consider only basket and chandelier interneurons, these cells fire out of phase compared with principal neurons during theta oscillations (4-8 Hz). Our data predict that these conditions would favour expression of CP-AMPAR-mediated LTP at both FF and FB afferents. Since FF inhibition has been shown to be critical to ensure the temporal fidelity of principal cell discharge [START_REF] Pouille | Enforcement of temporal fidelity in pyramidal cells by somatic feed-forward inhibition[END_REF], non-Hebbian LTP of FF inputs onto PV INs may then contribute to maintain this fidelity when excitatory inputs onto principal cells are potentiated [START_REF] Lamsa | Hebbian LTP in feed-forward inhibitory interneurons and the temporal fidelity of input discrimination[END_REF]. Conversely, firing of PV baskets cells is time locked with pyramidal cells during sharp waves ripples (SWRs; [START_REF] Klausberger | Neuronal diversity and temporal dynamics: the unity of hippocampal circuit operations[END_REF] which may contribute to memory consolidation [START_REF] Girardeau | Selective suppression of hippocampal ripples impairs spatial memory[END_REF]. During SWRs, large populations of CA3 and CA1 pyramidal neurons are entrained into brief but very high frequency oscillations . Our data predict that, although both FF and FB inputs onto PV baskets cells may be active during SWRs, FB inputs may undergo a selective potentiation through activation of NMDARs. Potentiated inputs may then contribute to improve spike timing precision of these cells during subsequent theta oscillations. In fact, specific ablation of NR1 in PV INs reduced theta and increased gamma oscillation power [START_REF] Korotkova | NMDA receptor ablation on parvalbumin-positive interneurons impairs hippocampal synchrony, spatial representations, and working memory[END_REF], an effect that remains unexplained mechanistically. It seems unlikely that NMDARs participate in phasic excitatory synaptic transmission onto PV INs. Instead, our data suggest NMDAR-dependent plasticity at FB inputs onto PV INs may be required to adjust their efficacy and maintain the precisely timed, clockwork control of PV INs over principal cell activity. Realistic models [START_REF] Traub | Contrasting roles of axonal (pyramidal cell) and dendritic (interneuron) electrical coupling in the generation of neuronal network oscillations[END_REF][START_REF] Volman | Downregulation of parvalbumin at cortical GABA synapses reduces network gamma oscillatory activity[END_REF]) will be needed to fully explore the functional consequences of input specific learning rules in various subtypes of hippocampal PV INs.

C 2013

 2013 The Authors. The Journal of Physiology C 2013 The Physiological Society contribution of NMDARs to EPSCs at FB vs. FF excitatory inputs onto PV INs may then reflect a similar difference in GluN2 subunit composition by comparing the effect of the specific GluN2B antagonist Ro25-6981. Application of 1 μM Ro25-6981 reduced NMDAR-mediated EPSCs to a similar extent at FB and FF inputs onto PV INs (-34 ± 12 vs.

Figure 1 .

 1 Figure 1. Postsynaptic receptor content at feed-forward vs. feedback excitatory inputs onto PV interneurons (INs) A, infrared differential interference contrast (left) and fluorescence (right) video images of a CA1 GFP-expressing interneuron in an acute hippocampal slice from PV Cre ::RCE mouse. Scale, 20 μm. B, evoked EPSCs recorded at +60 mV or -60 mV for a pyramidal neuron (Pyr) and PV IN (PV) in the presence of 100 μM APV. C and D, current-voltage relation (C) and rectification index (D) of AMPAR-mediated EPSCs recorded from pyramidal neurons (n = 4, grey) or PV INs (n = 7) upon stimulation of either FF (white ) or FB (black) afferents. E, time course of Naspm-induced partial block of EPSCs evoked in PV INs upon FF (white) or FB (black) afferent stimulation (n = 10). F, normalized EPSC amplitude upon 15-25 min of Naspm application. Mean data from 2 pyramidal neurons and 10 PV INs. G, left: 15 consecutive evoked EPSCs (grey) and mean currents (black) recorded at +60 mV or -60 mV for a pyramidal neuron (Pyr) and PV IN (PV). G, right: NMDA/AMPA ratio for EPSCs recorded from PV INs upon stimulation of FF (white) or FB (black) afferents (n = 12). H, left: NMDAR-mediated EPSCs recorded in the presence of 10 μM NBQX, before (grey) and after (black) application of Ro25-6981. H, right, normalized amplitude of the NMDA EPSC upon application of Ro25-6981 (n = 5). * P < 0.05; n.s., non-significant.

Figure 2 .

 2 Figure 2. Distinct plasticity rules at FF vs. FB excitatory synapses onto PV INs A-D, after a 5 min. control recording, afferent-specific stimulation (400 pulses at 5 Hz, arrowhead) was paired with either postsynaptic hyperpolarization to -90 mV (anti-Hebbian induction, A, B) or depolarization to 0 mV (Hebbian induction, C, D). Left, averaged EPSCs are shown for individual recordings before (grey) and after (black) pairing. Right, summary data from all experiments. Anti-Hebbian plasticity was observed at both FF (A, n = 10) and FB synapses (B, n = 9). However, Hebbian plasticity was induced only at FB (D, n = 8) but not FF (C, n = 13) synapses onto PV INs.

Figure 3 .

 3 Figure 3. Anti-Hebbian and Hebbian LTP in PV INs require activation of GluA2-lacking AMPARs and NMDARs, respectively A, application of Naspm (100 μM) immediately prior to pairing (arrowhead) prevented anti-Hebbian LTP at FB excitatory afferents (n = 6). Normalized EPSC amplitudes are shown for control (grey circles) and paired (black) pathways. No significant difference in the recovery of EPSCs from Naspm blockade was observed between control and paired pathways. B, C, application of 100 μM APV prevented the induction of Hebbian LTP at FB afferents (B, n = 11) without affecting induction of anti-Hebbian LTP at FF afferents (C, n = 14). D, summary data of all experiments showing the mean amplitude of EPSCs normalized to control upon Hebbian (0 mV) or anti-Hebbian (-90 mV) pairing in control conditions or in the presence of either Naspm or APV for FF (white) or FB (black) afferents. Changes in EPSC amplitude upon pairing are compared in the presence of antagonists vs. control conditions. * P < 0.05; n.s., non-significant.

Figure 4 .

 4 Figure 4. Presynaptic function is unaffected upon Hebbian and anti-Hebbian LTP in PV INs A, averaged EPSCs evoked by paired pulses (50 ms apart) before (grey) and after (black) pairing-induced LTP. Top, anti-Hebbian LTP of FF inputs. Bottom, Hebbian pairing of FB inputs. Right, the second EPSC (EPSC2) is shown before and after pairing, after normalization to the peak amplitude of the first (EPSC1 scaled). No change in PPR was detectable in either condition. B, normalized PPR for control pathway or potentiated pathway. 5 Hz afferent stimulation was paired with hyperpolarization to -90 mV (FF inputs, white circles, n = 15; FB inputs, black circles, n = 14) or depolarization to 0 mV (FB inputs; black circles, n = 10). C, left: A1 receptor antagonist DPCPX (100 nM) increased EPSC amplitude in PV INs by about 40% (n = 10). C, right: compared effect of Naspm on evoked EPSCs in control conditions (grey circles, n = 17) or after potentiation by DPCPX (n = 10, black circles). EPSC amplitudes were normalized to mean values prior to Naspm application. The time course of Naspm-induced blockade of EPSCs (fitted to a single exponential) was faster in PV INs recorded in the presence of DPCPX. * P < 0.05. D, summary data of the decay time constant (tau) of EPSC blockade by Naspm in either condition. E, F, similar experiment in which the effect of Naspm was monitored on EPSCs evoked from two independent FF pathways (paired, black circles; control, white circles). Arrowhead indicates pairing. EPSC amplitudes were normalized to values prior to Naspm application. No significant difference in the decay of EPSC amplitude was detected between the two pathways (n = 10).

Figure 5 .

 5 Figure 5. Postsynaptic Hebbian and anti-Hebbian LTP in PV INs A, EPSCs evoked by focal uncaging of MNI-Glutamate onto proximal dendrites of PV INs in st. radiatum. Average of 30 uEPSCs normalized in amplitude and compared to 200 averaged mEPSCs recorded from the same cell showing fast onset kinetics despite slower decay of uEPSCs as compared to mEPSCs. B, right: uEPSCs evoked at various positions along a PV IN apical dendrite in st. radiatum. 10 individual uEPSCs (grey) are shown superimposed with averaged current (black) at each position. B, left, graph showing peak amplitude and decay time constant (decay tau) as a function of distance from the spot of maximal amplitude and minimal decay for the cell shown in right panel. Note the sharp decrease in amplitude and increase in decay kinetics of uEPSCs when light spot is moved away from optimal position. C, pairing consisting of 100 uEPSCs at 5 Hz with membrane hyperpolarization to -90 mV (arrowhead) leads to a persistent potentiation of uEPSC amplitude (white circles, n = 8 cells) as compared to control uEPSCs (grey circles, n = 3 cells). D, averaged uEPSCs from one such experiment before (n = 30) and 10-20 min after (n = 40) pairing. E, similar experiment on uEPSCs evoked by glutamate uncaging onto PV IN dendrites in st. oriens. Pairing of 100 uEPSCs at 5 Hz with membrane voltage clamped at 0 mV (arrowhead) leads to a potentiation of uEPSCs evoked at distal (>50 μm from soma close to alveus, black circles, n = 7 cells) but not proximal dendritic sites (20-40 μm from soma, black triangles, n = 6 cells) as compared to control (grey circles, n = 3 cells). F, averaged uEPSCs from one experiment with distally evoked uEPSCs before (n = 30) and 10-15 min after (n = 40) pairing.

Figure 6 .

 6 Figure 6. Frequency-dependent plasticity at FF vs. FB inputs onto PV INs A-D, long term plasticity induced by repetitive afferent stimulation (arrowheads) to either FF (white) or FB (black) inputs onto PV INs. 900 pulses were delivered at 1 (A), 5 (B) or 20 (C) Hz. In D, 100 pulses were delivered 5 times at 100 Hz while recording PV INs in current-clamp mode. Lower graphs represent averages of EPSC amplitudes recorded in voltage-clamp mode at -60 mV, normalized to control prior to conditioning stimulus; n = 5-7 cells for each frequency and each pathway. Upper graphs show sample recordings during conditioning stimulus at each frequency. Insets: detail of the first 150 ms. Note that 100 Hz stimulation only was sometimes sufficient to depolarize PV INs above firing threshold (action potentials are clipped due to filtering). E, 100 Hz stimulation was delivered to FB inputs in the presence of 100 μM APV to block NMDARs; n = 7 cells. F, 100 Hz stimulation was delivered to FF inputs while holding the postsynaptic cell to -90 mV in voltage-clamp (VC) mode to allow for activation of Ca 2+ -permeable AMPARs; n = 6 cells. G, summary of all experiments with afferent stimulations delivered at various frequencies (1, 5, 20 or 100 Hz) at FF (white) or FB (black) inputs onto PV INs, or FF inputs onto pyramidal neurons (grey). Each data point represents the mean ± SEM of normalized EPSC amplitudes measured between 10 and 20 min after conditioning stimulus. Note that beyond 20 Hz stimulation, FF inputs onto PV INs show no LTP, in contrast to FB inputs and FF inputs onto pyramidal cells. # P < 0.05 FF inputs onto pyramidal cells compared to FF and FB inputs onto PV INs. * P < 0.05 FF compared to FB inputs onto PV INs.

C 2013 The Authors. The Journal of Physiology C 2013 The Physiological Society

J Physiol 00.00

N. Le Roux and others J Physiol 00.00

Acknowledgements

We thank S. Arber and G. Fishell for kindly providing Pvalb tm1(cre)Arbr and RCE:LoxP mice, T. Irinopoulou for assistance with imaging, M. Mameli for helpful discussions and R. Miles and M. Mameli for critical reading of the manuscript. This work was supported by the Institut National de la Santé et de la Recherche Médicale (Avenir program grant to J.C.P.), Ecole des Neurosciences de Paris (fellowship to U.L.B.), research grants (to J.C.P.) from the City of Paris, the Fyssen Foundation and the Agence Nationale de la Recherche. The authors declare no conflict of interest.