Stuttering Interneurons Generate Fast and Robust Inhibition onto Projection Neurons with Low Capacity of Short Term Modulation in Mouse Lateral Amygdala

et al. (2013) Stuttering Interneurons Generate Fast and Robust Inhibition onto Projection Neurons with Low Capacity of Short Term Modulation in Mouse Lateral Amygdala. PLoS ONE 8(3): e60154. doi:10.1371/journal.pone.0060154 Stuttering Interneurons Generate Fast and Robust Inhibition onto Projection Neurons with Low Capacity of Short Term Modulation in Mouse Lateral Amygdala Xing Pan 0 Chen Song 0 Xiao-Bin Xu 0 Ye He 0 Zhi-Peng Liu 0 Min Wang 0 Xin Zhang 0 Bao-Ming Li 0 Bing- 0 Lin Mei, Georgia Regents University, United States of America 0 1 Laboratory of Fear and Anxiety Disorder, Institute of Life Science, Nanchang University , Nanchang , China , 2 Department of Pharmacology, Nanchang University , Nanchang , China The stuttering interneurons (STi) represent one minor subset of interneuron population and exhibit characteristic stuttering firing upon depolarization current injection. While it has been long held that the GABAergic inhibitory transmission largely varies with the subtype identity of presynaptic interneurons, whether such a rule also applies to STi is largely unknown. Here, by paired recording of interneuron and their neighboring projection neuron in lateral amygdala, we found that relative to the fast spiking and late spiking interneurons, the STi-evoked unitary postsynaptic currents onto the projection neurons had markedly larger amplitude, shorter onset latency and faster rising and decay kinetics. The quantal content and the number of vesicles in the readily releasable pool were also larger in synapses made by STi versus other interneurons. Moreover, the short-term plasticity, as reflected by the paired pulse depression and depolarization-induced suppression of inhibition, was the least prominent in the output synapses of STi. Thus, the fast and robust inhibition together with its low capacity of short term modulation may suggest an important role for STi in preventing the overexcitation of the projection neurons and thus gating the information traffic in amygdala. - The lateral nucleus of amygdala (LA), a gatekeeper of the multimodal sensory information from cortical and subcortical areas entering the amygdala, has been generally recognized to play a critical role in the acquisition, storage and expression of emotional information such as fear and anxiety [13]. Whereas the excitatory projection neurons (PNs) mediate the signal transfer between LA and its down- and upstream brain areas [46], the local GABAergic interneurons (INs) prevent the overexcitation of PNs and ensure the appropriate expression of fear and anxiety through establishing the highly inhibitory tone in amygdala [7,8]. The impairment in amygdala inhibition closely correlates with the development of a series of mental disorders such as posttraumatic stress disorders [9,10]. As in hippocampus and cortex, the INs in LA exhibit wide diversity in terms of their morphological, neurochemical and electrophysiological features [11]. Studies using immunostaining of their molecular markers have revealed some major non-overlapping subtypes of INs in LA with each expressing parvalbumin (PV), cholecystokinin (CCK) or somatostatin (SOM) [1214]. Based on their spiking response to the current step injection, the INs can be classified into multiple sets such as fast spiking interneurons (FSi), late spiking interneurons (LSi), accommodating INs (ACi) and STi [1518]. Of these, the STi constitute a minor subset of the IN population and are characterized by bursts of action potentials intermingled with variable quiescent periods upon the sustained depolarization current injection. Accumulating evidence has shown that the inhibition imposed on the target neurons largely depend on the subtype identity of the presynaptic Ins [19,20]. For example, in hippocampus, the fastspiking basket cells generate fast and strong perisomatic inhibition onto the PNs, the late-spiking neurogliaform cells, on the other hand, provide slow and weak inhibitory signal through the connections distal to the soma [21]. Although the STi have been identified in multiple brain areas such as cortex, striatum and amygdale [15,16,22,23], very little is known about the properties of the unitary inhibitory transmission mediated by STi. It is yet unclear whether the rule also applies to the STi that presynaptic INs dictate the inhibitory transmission. Specifically, do the STi, which fire in a pattern clearly distinguishable from other INs, also generate unique form of inhibition onto their nearby PNs? To answer this, we made simultaneous recording of IN-PN pairs from GAD-67 GFP knock-in mice and compared the properties of unitary inhibitory postsynaptic currents (uIPSCs) in connections made by STi and other IN subtypes. We found that relative to other INs, the STi evoked faster and more robust inhibition onto Figure 1. The uIPSCs evoked by different subtypes of INs onto PNs in LA. A, Schematic graph showing the simultaneous recording of IN-PN pairs in LA. B, Representative traces showing the firing patterns of STi, FSi, LSi and ACi upon injection of 1 s threshold current (middle trace) and current 80pA above the threshold (top trace). The bottom shows the pattern of current injection onto INs. C, Representative traces showing the uIPSCs in individual PNs evoked by distinct subtypes of INs. Insets show the expanded graph and the dashed lines indicate the peak of spike in INs (left) and the onset of uIPSCs in PNs (right). doi:10.1371/journal.pone.0060154.g001 nearby PNs. Moreover, the short-term prominent in the output synapses of STi. Materials and Methods plasticity was less Slice preparation All experimental procedures involving animals were approved by the Animal Ethics Committee of Nanchang University. Amygdala slices were prepared as previously described from 4 5 weeks old heterozygous GAD67-GFP(Dneo) male mice in which GFP is selectively expressed in Ins [24,25]. Briefly, mice were sacrificed by decapitation and brains were quickly removed to icecold oxygenated (95% O2/5% CO2) artificial cerebrospinal fluid (ACSF) containing (in mM): 124 NaCl, 2.5 KCl, 1 MgSO4, 2.5 CaCl2, 10 glucose, and 26 NaHCO3 (pH 7.30). Slices containing LA of about 350 mm were cut with a Leica VT 1000S tissue slicer and maintained at room-temperature for at least one hour before recording. Electrophysiological Recording Slices were transferred to a recording chamber continuously superfused with ACSF at a constant rate of about 60 ml/h and the recording temperature was held at 2961uC. Dual whole cell recordings were performed in IN-PN pairs in LA with an EPC-10 amplifier and Patchmaster software (HEKA Elektronik, Germany). The PNs were visualized under guidance of DIC/infrared optics and the INs by their green fluorescence. Data were filtered at 2 K Hz using the patch-clamp amplifier circuitry and digitized at 10 k Hz. The patch pipettes for recording PNs were filled with (in mM): 100 CsCl, 20 Cs-methanesulfonate, 5 NaCl, 2 MgCl2, 10 HEPES, and 0.2 EGT (...truncated)