GABAergic inhibitory interneurons constitute a population of hippocampal cells whose high degree of anatomical and functional divergence make them suitable candidates for controlling the activity of large populations of principal neurons. GABAergic inhibitory interneurons play a major role in the synchronization of neuronal activity and are involved in the generation of large-scale network oscillations. Thus, interneurons function as a clock that dictates when principal cells fire during suprathreshold excitatory drive. Interneurons receive strong excitatory glutamatergic innervation via numerous anatomically distinct afferent projections, and it was recently demonstrated that the molecular composition of both classes of the AMPA-preferring glutamate receptors expressed at interneuron synapses are often distinct from those found at principal cell synapses. Furthermore, single inhibitory interneurons can synthesize distinct AMPA receptors with defined subunit composition and target them to synaptic domains innervated by different afferent inputs. Using high-resolution whole-cell patch clamp recording techniques in brain slices of hippocampus and the auditory and barrel cortices, we investigate differential mechanisms of synaptic transmission onto hippocampal inhibitory interneurons and the role of intrinsic voltage-gated channels in regulating interneuron excitability.
Mossy fiber synaptic transmission: control by the metabotropic glutamate receptor 7 (mGluR7) of the bidirectional plasticity of mossy fiber–stratum lucidum feedforward inhibition
Pelkey; in collaboration with Lacaille, Racca, Roche
Activity-dependent alterations in synaptic efficacy are considered the cellular substrate that underlies learning and memory formation. As such, the alterations have been implicated in neurodegenerative disorders such as epilepsy and chronic pain. Despite rapid progress in elucidating cellular mechanisms responsible for long-term plasticity at excitatory synapses between principal cells, relatively little insight has been gained into the plasticity of excitatory transmission onto inhibitory interneurons. Indeed, bidirectional plasticity of excitatory drive onto any identified interneuron population has not been previously observed. Moreover, excitatory transmission typically displays cell target–specific regulation, indicating that the rules governing plasticity at principal cell synapses cannot be applied to inputs onto interneurons. For instance, hippocampal mossy fiber (MF) inputs to CA3 stratum lucidum interneurons (SLINs) undergo long-term depression (LTD) following high-frequency stimulation (HFS), in contrast to MF-pyramid (PYR) synapses, where long-term potentiation (LTP) occurs. Furthermore, activity-induced potentiation of MF-SLIN transmission has not been previously observed. We have now shown that the presynaptically located metabotropic glutamate receptor subtype 7 (mGluR7), whose activation and surface expression governs the direction of plasticity, is a metaplastic switch at MF-SLIN synapses. In naive slices, mGluR7 activation during high-frequency stimulation generates MF-SLIN LTD, depressing presynaptic release through a PKC-dependent mechanism. Moreover, following agonist exposure, mGluR7 undergoes internalization, unmasking the ability of MF-SLIN synapses to undergo presynaptic potentiation in response to the HFS that induces LTD in naive slices. Thus, selective mGluR7 targeting to MF terminals contacting SLINs and not PYRs provides cell target–specific plasticity and bidirectional control of feedforward inhibition.
Using two-photon Ca2+ imaging of MF terminals in acute mouse hippocampal slices, we next examined the potential role of mGluRs in the regulation of presynaptic Ca2+ transients (CTs) evoked by MF stimulation. The group II mGluR agonist DCGIV depressed CTs equally in large MF boutons (MFBs) onto pyramidal cells and smaller MF filopodia (MFF) onto inhibitory interneurons. In contrast, the distinct MF synapses displayed different sensitivities to the group III mGluR agonist L-AP4. At concentrations saturating for mGluR4/8/6, L-AP4 reversibly inhibited MFB CTs without altering MFF CTs. At concentrations sufficient to recruit mGluR7, L-AP4 still reversibly depressed MFB CTs but also depressed MFF CTs. Moreover, depression at MFF terminals was irreversible (LTD of CTs) if L-AP4 was applied with continuous stimulation of MF afferents, but reversible if stimulation was interrupted during agonist perfusion. These observations are consistent with our findings that L-AP4 reversibly depresses MF-CA3 pyramid excitatory post-synaptic currents (EPSC)s at all concentrations tested and produces LTD of MF-SLIN EPSCs only at high concentrations, with no effect at low concentrations. Our findings solidify the hypothesis that distinct mGluRs localize to MFBs and MFFs and mediate short- and long-term inhibition of MF-PYR and MF-SLIN synapses and indicate that mGluR7-dependent MF-SLIN LTD involves a long-term depression of presynaptic Ca2+ transients, presumably reducing the release probability of neurotransmitter release.
Lawrence JJ, Grinspan ZM, McBain CJ. Quantal analysis at mossy fiber targets in the CA3 region of the rat hippocampus. J Physiol 2004;554:175-193.
Lawrence JJ, McBain CJ. Containing the detonation: feedforward inhibition in the CA3 hippocampus. Trends Neurosci 2003;26:631-640.
Lei S, McBain CJ. Two loci of expression for long term depression at hippocampal mossy fiber-interneuron synapses. J Neurosci 2004;24:2112-2121.
Pelkey KA, Lavezzari G, Racca C, Roche KW, McBain CJ. mGluR7 is a metaplastic switch controlling bi-directional plasticity of feedforward inhibition. Neuron 2005;46:89-102.
Pelkey KA, McBain CJ. How to dismantle a detonator synapse. Neuron 2005;45:327-329.
Depolarization-induced long-term depression at hippocampal mossy fiber–CA3 pyramidal neuron synapses
Ho, Pelkey; in collaboration with Huganir
Classically, long-term plasticity at mossy fiber–CA3 pyramid synapses (MF-CA3) is considered to result from alterations in presynaptic release via cAMP/PKA-dependent signaling cascades. However, depolarization of CA3 pyramids in young animals (between P6 and P20) induces a persistent depression (DID) of MF-CA3 transmission attributable to decreased postsynaptic AMPAR function. This novel form of long-term depression is independent of NMDARs, mGluRs, cannabinoid receptors, opioid receptors, or coincident synaptic activity but is dependent on postsynaptic Ca2+ elevation through L-type Ca2+ channels and release from InsP3 receptor–sensitive intracellular stores. Ca2+ imaging of both proximal and distal CA3 pyramidal neuron dendrites demonstrated that the depolarizing induction paradigm differentially elevated intracellular Ca2+ levels. We observed L-type Ca2+ channel activation only at the most proximal locations at which MFs make synapses. Depolarization-induced LTD did not occlude the conventional 1 Hz–induced LTD or vice versa, suggesting that independent mechanisms underlie each form of plasticity. The paired-pulse ratio and coefficient of variation of synaptic transmission were unchanged after LTD induction, suggesting that the expression locus of LTD is postsynaptic. Moreover, peak-scaled nonstationary variance analysis indicated that depolarization-induced LTD correlated with a reduction in postsynaptic AMPA receptor numbers without a change in AMPA receptor conductance.
More recently, we examined the molecular details of DID, focusing on the scaffold protein PICK1 and the kinase PKC, given their importance in other post-synaptically expressed LTDs. In PICK1 knockout mice (–/–), EPSCs remained similar to those of controls 5 minutes following the DID induction paradigm. In addition, the failure of EPSCs to depress after DID induction in the presence of PKC inhibitors suggests a role for PKC in DID. Emerging evidence indicates that PICK1 and PKC control surface GluR2 expression, thereby regulating AMPAR subunit composition. Thus, we next examined whether GluR2-lacking, Ca2+-permeable AMPARs (CP-AMPARs) participate in MF-CA3 transmission. Surprisingly, CP-AMPARs contributed significantly to MF-CA3 transmission in PICK1+/+ slices as determined by inwardly rectifying I-V relations of MF-CA3 EPSCs. In contrast, MF-CA3 synapses in PICK1–/– slices yielded significantly more linear I-V relations, indicating that GluR2-containing AMPARs dominate MF-CA3 transmission in the absence of PICK1. Our working hypothesis posits that depolarization-induced depression of mossy fiber–pyramidal cell transmission results from conversion of GluR2-lacking MF-CA3 synapses to those containing GluR2 in a PICK1- and PKC-dependent manner.
Lei S, Pelkey K, Topolnik L, Congar P, Lacaille J-C, McBain CJ. Depolarization-induced long term depression at hippocampal mossy fiber-CA3 pyramidal neuron synapses. J Neurosci 2003;23:9786-9795.
Muscarinic control of spike timing and reliability in hippocampal interneurons
Lawrence
Hippocampal theta oscillations are thought to be the primary means by which neuronal assemblies bind together distinct aspects of spatial experience. The activation of hippocampal interneurons by cholinergic afferents of the septum may play an important role in the initiation of theta oscillations. Recently, several interneuron subclasses have been shown to discharge at different times during the theta cycle; their discharge patterns are related to both the nature of their reciprocal connectivity with pyramidal cells and the intrinsic membrane properties of each interneuron class. Although cholinergic neuromodulation has been shown to have potent effects on the resting membrane potential of different interneuron types, the impact of cholinergic neuromodulation on spike timing and reliability is unknown. We focused on a physiologically and anatomically well-described interneuron subtype, the stratum oriens/lacunosum-moleculare (O-LM) interneurons. These interneurons are poised to receive cholinergic input from the medial septum and to modulate the flow of information from the entorhinal cortex to the hippocampus.
We find that muscarinic receptor activation consistently and potently enhanced the excitability of O-LM interneurons and produced large, sustained afterdepolarizations in response to depolarizing current injection. The afterdepolarization was mediated by the combined activation of M1 and M3 muscarinic receptors and was triggered in part by the downregulation of two potassium conductances, an M-current and a calcium-dependent afterhyperpolarization current, and the emergence of a calcium-dependent cationic conductance. To assess how muscarine influences the oscillatory properties of O-LM interneurons, we introduced sinusoidal current injections at a variety of frequencies. Muscarine caused an increase in action potential reliability, a shift in action potential timing to earlier in the cycle, and an increase in spike timing precision. Interestingly, the enhanced spike reliability and increased spike timing precision was approximately limited to theta frequencies, suggesting that cholinergic neuromodulation tunes the intrinsic membrane properties of O-LM interneurons to match the frequency of extrinsic synaptic input. Such post-synaptic specialization would allow cholinergic afferents from the medial septum fine control over the degree of engagement of O-LM interneurons in network activity.
Differential distribution of KCNQ/M channel subunits in hippocampal CA1 interneurons
Lawrence, Churchill, Travis; in collaboration with Saraga, Skinner
Potassium channels of the KCNQ subtype are thought to underlie the macroscopic M-current (IM). Historically, M-currents have been shown to play a key role in controlling action potential generation and firing pattern. While such conductance has been well described in principal cells, it is unclear whether interneurons also possess IM. We have taken a combined electrophysiological, immunocytochemical, and computational approach to characterize IM in CA1 stratum oriens/alveus (OA) interneurons.
Using patch clamp techniques from acute mouse slices, we first determined whether IM was present in visually identified hippocampal CA1 OA interneurons. We had shown previously that OA interneurons possess both transient and sustained outward currents, with the latter thought to be mediated predominantly by delayed rectifier (Kv3) potassium channels. However, selective IM antagonists linopirdine and XE991 removed about 30 percent of the total sustained outward current present at +40mV, suggesting a significant contribution of KCNQ/M current. Test pulses between –60 and +40 mV revealed a linopirdine-sensitive (LS) current component. An analysis of the time course of deactivation disclosed that the KCNQ/M current possessed an intermediately decaying component of the outward current while leaving both faster- and slower-decaying components unchanged. This kinetic profile suggests that M-currents may contribute considerably to the intermediate component of afterhyperpolarization. The currents’ voltage-dependence, kinetics, and pharmacology all strongly suggest that KCNQ2 channels mediate the KCNQ/M conductance in OA interneurons. We also found that blocking IM with linopirdine affected cell excitability by causing an increase in action potential firing.
Immunohistochemistry performed in GAD67-eGFP transgenic mice, which selectively label somatostatin-positive interneurons (a population that includes the O-LM cell), reveals a somatodendritic localization of KCNQ2/3 subunits. Consistent with the electrophysiological IM data on OA interneurons, about 40 percent of GFP-positive OA interneurons are KCNQ2-positive. Interestingly, GFP-positive stratum radiatum interneurons express less KCNQ2. A similar distribution is found for KCNQ3 subunits. Using these quantitative characteristics, we have now incorporated IKCNQ/M into a multicompartmental O-LM interneuron model that is both physiologically and morphologically realistic. Using the model to explore different cellular IKCNQ/M distributions and densities, we find that somatodendritic placement of KCNQ/M channels best describes experimentally measured IM in OA interneurons.
Recruitment of a network of hippocampal stratum lucidum interneurons by a frequency-dependent switch of synaptic inhibition polarity
Banke
Within the hippocampus, feedforward inhibitory drive provides a potent braking mechanism of principal cell excitation. We were interested in determining the impact of high-frequency synaptic inhibition among SLINs, in particular whether a frequency-dependent switch in polarity occurred that could entrain SLINs to provide coordinated feedforward inhibition.
We recorded from CA3 SLINs in the perforated patch mode to preserve intracellular chloride homeostasis. Stimulation of inhibitory afferents at 40 to 100Hz caused a rapid reversal of inhibition in at least two populations of SLINs (Class I and II) such that inhibitory synaptic potentials depolarized and were capable of driving action potential firing. The duration of the switch was brief, and full recovery occurred within 2 to 3 seconds. Depolarizing inhibitory post-synaptic potentials (IPSPs) deviated from the control current-voltage relationship, suggesting a temporary redistribution of Cl– gradients involving both KCC2 and NKCC1. GABAA receptor antagonists blocked the polarity switch in Class I SLINs. In Class II SLINs, in addition to a reversal of the Cl– gradient, we observed a large depolarizing envelope that was blocked by mGluR group I/II antagonists. To investigate the impact of the switch in inhibitory drive on the hippocampal network, we recorded spontaneous IPSCs (inhibiting postsynaptic currents) from pyramidal cells. With excitation blocked, pyramidal cells showed a clear three-fold increase in sIPSC frequency immediately after the high-frequency stimulation and a return of the frequency to control levels within 2 seconds, a time course consistent with the observed polarity switch on interneurons. The sIPSPs increased transiently but effectively blocked the ability of pyramidal cells to fire repetitively and were blocked by bicuculine or inclusion of the Na-channel blocker TTX. The data demonstrate the existence of a brief window following high-frequency afferent activity during which GABAA-mediated synaptic events depolarize CA3 SLINs, ensuring a brief but coordinated feedforward inhibitory drive onto CA3 pyramidal cells.
Mediation of metabotropic regulation of the slow and medium AHP currents in CA3 pyramidal cells by the kainate receptor subunit GluR6
Fisahn1; in collaboration with Heinemann
Kainate is a well-known excitotoxic that increases excitability and generates seizure-like activity as well as gamma oscillations in the hippocampus. Changes in excitability have consequences for the action potential firing characteristics of neurons, which in turn are important for the overall behavior and output of neuronal networks such as synchronized oscillations or seizure-like activity. Prolonged depolarization of neurons above their firing threshold leads to initially sustained generation of action potentials. In most principal neuron types in the hippocampus, however, the generation of action potentials slows down and stops before the depolarizing stimulus has abated. This spike frequency accommodation is brought about by a number of conductances, including a calcium-activated potassium current with slow decay time that hyperpolarizes the cell membrane (IsAHP). The slow after hyperpolarization (AHP) current therefore governs the length and frequency of bursts of action potentials. Likewise, the length and frequency of single action potentials is influenced by calcium-activated hyperpolarizing potassium currents, which in pyramidal neurons have medium and fast decay times (ImAHP and IfAHP). A recent study showed that direct activation of kainate receptors (KARs) on CA1 pyramidal cells decreases IsAHP. Moreover, the action of kainate was shown to be metabotropic, activating a PKC-based signaling pathway. We were interested in investigating the role of different KAR subunits in modulating IsAHP and ImAHP and their influence on repetitive action potential generation. To that end, we used whole-cell patch clamp recordings from CA3 pyramidal cells of wild-type and KAR knockout mice to investigate the role of KAR subunits in the metabotropic regulation of the slow and medium afterhyperpolarization currents (IsAHP and ImAHP). The kainate-induced decrease in IsAHP and ImAHP amplitude was PKC-dependent and absent from GluR6–/– but not from GluR5–/– neurons. Our findings suggest that activation of GluR6-containing KARs modulates AHP amplitude and influences the firing frequency of pyramidal neurons.
Fisahn A, Heinemann SF, McBain CJ. The kainate receptor subunit GluR6 mediates metabotropic regulation of the slow and medium AHP currents in mouse hippocampal neurons. J Physiol 2005;562:183-198.
1André Fisahn, PhD, former Visiting Fellow, now at the Karolinska Institute, Stockholm, Sweden
Collaborators
Stephen Heinemann, PhD, The Salk Institute, La Jolla, CA
Richard Huganir, PhD, Howard Hughes Medical Institute, The Johns Hopkins University School of Medicine, Baltimore, MD
Jean-Claude Lacaille, PhD, University of Montreal, Montreal, Canada
Claudia Racca, PhD, University of Leeds, Leeds, UK
Katherine Roche, PhD, Basic Neurosciences Program, NINDS, Bethesda, MD
Fernanda Saraga, PhD, University Toronto, Canada
Frances Skinner, PhD, University of Toronto, Toronto, Canada
For further information, contact mcbainc@mail.nih.gov.