Arnaud Ruiz

Monosynaptic GABAergic Signaling from Dentate to CA3 with a Pharmacological and Physiological Profile Typical of Mossy Fiber Synapses

Mossy fibers are the sole excitatory projection from dentate gyrus granule cells to the hippocampus, where they release glutamate, dynorphin, and zinc. In addition, mossy fiber terminals show intense immunoreactivity for the inhibitory neurotransmitter GABA. Fast inhibitory transmission at mossy fiber synapses, however, has not previously been reported. Here, we show that electrical or chemical stimuli that recruit dentate granule cells elicit monosynaptic GABA(A) receptor-mediated synaptic signals in CA3 pyramidal neurons. These inhibitory signals satisfy the criteria that distinguish mossy fiber-CA3 synapses: high sensitivity to metabotropic glutamate receptor agonists, facilitation during repetitive stimulation, and NMDA receptor-independent long-term potentiation. GABAergic transmission from the dentate gyrus to CA3 has major implications not only for information flow into the hippocampus but also for developmental and pathological processes involving the hippocampus.

GABAA Receptors at Hippocampal Mossy Fibers

Presynaptic GABAA receptors modulate synaptic transmission in several areas of the CNS but are not known to have this action in the cerebral cortex. We report that GABAA receptor activation reduces hippocampal mossy fibers excitability but has the opposite effect when intracellular Cl- is experimentally elevated. Synaptically released GABA mimics the effect of exogenous agonists. GABAA receptors modulating axonal excitability are tonically active in the absence of evoked GABA release or exogenous agonist application. Presynaptic action potential-dependent Ca2+ transients in individual mossy fiber varicosities exhibit a biphasic dependence on membrane potential and are altered by GABAA receptors. Antibodies against the alpha2 subunit of GABAA receptors stain mossy fibers. Axonal GABAA receptors thus play a potentially important role in tonic and activity-dependent heterosynaptic modulation of information flow to the hippocampus.

Presynaptic GABAA receptors enhance transmission and LTP induction at hippocampal mossy fiber synapses

Presynaptic GABAA receptors (GABAARs) occur at hippocampal mossy fiber synapses. Whether and how they modulate orthodromic signaling to postsynaptic targets is poorly understood. We found that an endogenous neurosteroid that is selective for high-affinity δ subunit–containing GABAARs depolarized rat mossy fiber boutons, enhanced action potential–dependent Ca2+ transients and facilitated glutamatergic transmission to pyramidal neurons. Conversely, blocking GABAARs hyperpolarized mossy fiber boutons, increased their input resistance, decreased spike width and attenuated action potential–dependent presynaptic Ca2+ transients, indicating that a subset of presynaptic GABA receptors are tonically active. Blocking GABAARs also interfered with the induction of long-term potentiation at mossy fiber–CA3 synapses. Presynaptic GABAARs therefore facilitate information flow to the hippocampus both directly and by enhancing LTP.

GABA and GABAA receptors at hippocampal mossy fibre synapses

Anatomical and electrophysiological evidence has raised the possibility that corelease of GABA and glutamate occurs at hippocampal mossy fibre synapses which, however, lack the vesicular GABA transporter VGAT. Here, we apply immunogold cytochemistry to show that GABA, like glutamate, has a close spatial relation to synaptic vesicles in rat mossy fibre terminals, implying that a mechanism exists to package GABA in synaptic vesicles. We also show that GABAA and AMPA receptors are colocalized at mossy fibre synapses. The expression of GABA and GABAA receptors is, however, weaker than in inhibitory synapses. Electrical stimuli that recruit mossy fibres evoke monosynaptic GABAA receptor‐mediated signals in post‐synaptic targets that show marked frequency‐dependent facilitation and sensitivity to group II metabotropic receptors, two features that are characteristic of mossy fibre transmission. These results provide further evidence for GABA and glutamate cotransmission at mossy fibre synapses, although paired pre‐ and post‐synaptic recordings will be required to determine the role of GABA at this unusual synapse.

Analog Modulation of Mossy Fiber Transmission Is Uncoupled from Changes in Presynaptic Ca2

Subthreshold somatic depolarization has been shown recently to modulate presynaptic neurotransmitter release in cortical neurons. To understand the mechanisms underlying this mode of signaling in the axons of dentate granule cells (hippocampal mossy fibers), we have combined two-photon Ca2+ imaging with dual-patch recordings from somata and giant boutons forming synapses on CA3 pyramidal cells. In intact axons, subthreshold depolarization propagates both orthodromically and antidromically, with an estimated length constant of 200–600 μm depending on the signal waveform. Surprisingly, presynaptic depolarization sufficient to enhance glutamate release at mossy fiber–CA3 pyramidal cell synapses has no detectable effect on either basal Ca2+-dependent fluorescence or action-potential-evoked fluorescence transients in giant boutons. We further estimate that neurotransmitter release varies with presynaptic Ca2+ entry with a 2.5-power relationship and that depolarization-induced synaptic facilitation remains intact in the presence of high-affinity presynaptic Ca2+ buffers or after blockade of local Ca2+ stores. We conclude that depolarization-dependent modulation of transmission at these boutons does not rely on changes in presynaptic Ca2+.

Mutations in SLC12A5 in epilepsy of infancy with migrating focal seizures.

The potassium-chloride co-transporter KCC2, encoded by SLC12A5, plays a fundamental role in fast synaptic inhibition by maintaining a hyperpolarizing gradient for chloride ions. KCC2 dysfunction has been implicated in human epilepsy, but to date, no monogenic KCC2-related epilepsy disorders have been described. Here we show recessive loss-of-function SLC12A5 mutations in patients with a severe infantile-onset pharmacoresistant epilepsy syndrome, epilepsy of infancy with migrating focal seizures (EIMFS). Decreased KCC2 surface expression, reduced protein glycosylation and impaired chloride extrusion contribute to loss of KCC2 activity, thereby impairing normal synaptic inhibition and promoting neuronal excitability in this early-onset epileptic encephalopathy.

A delayed response enhancement during hippocampal presynaptic plasticity in mice

High frequency afferent stimulation of chemical synapses often induces short‐term increases in synaptic efficacy, due to increased release probability and/or increased supply of readily releasable synaptic vesicles. This may be followed by synaptic depression, often caused by vesicle depletion. We here describe an additional, novel type of delayed and transient response enhancement phase which occurred during prolonged stimulation at 5–20 Hz frequency of excitatory glutamatergic synapses in slices from the adult mouse CA1 hippocampal region. This second enhancement phase, which was most clearly defined at physiological temperatures and essentially absent at 24°C, was dependent on the presence of F‐actin filaments and synapsins I and/or II, and could not be ascribed to changes in presynaptic action potentials, inhibitory neurotransmission or glutamate receptor desensitization. Time course studies showed that the delayed response phase interrupted the synaptic decay 3–4 s after stimulus train initiation and continued, when examined at 5–10 Hz frequencies, for approximately 75 stimuli before decay. The novel response enhancement, probably deriving from a restricted pool of synaptic vesicles, may allow maintenance of synaptic efficacy during prolonged periods of excitatory synaptic activity.

Do mossy fibers release GABA

 Purpose: Mossy fibers are the sole excitatory projection from dentate gyrus granule cells to the hippocampus, forming part of the trisynaptic hippocampal circuit. They undergo signficiant plasticity during epileptogenesis and have been implicated in seizure generation. Mossy fibers are a highly unusual projection in the mammalian brain; in addition to glutamate, they release adenosine, dynorphin, zinc, and possibly other peptides. Mossy fiber terminals also show intense immunoreactivity for the inhibitory neurotrasnmitter γ‐aminobutyric acid (GABA), and immunoreactivity for GAD67. The purpose of this review is to present physiologic evidence of GABA release by mossy fibers and its modulation by epileptic activity.

Ionotropic receptors at hippocampal mossy fibers: roles in axonal excitability, synaptic transmission, and plasticity.

Dentate granule cells process information from the enthorinal cortex en route to the hippocampus proper. These neurons have a very negative resting membrane potential and are relatively silent in the slice preparation. They are also subject to strong feed-forward inhibition. Their unmyelinated axon or mossy fiber ramifies extensively in the hilus and projects to stratum lucidum where it makes giant en-passant boutons with CA3 pyramidal neurons. There is compelling evidence that mossy fiber boutons express presynaptic GABAA receptors, which are commonly found in granule cell dendrites. There is also suggestive evidence for the presence of other ionotropic receptors, including glycine, NMDA, and kainate receptors, in mossy fiber boutons. These presynaptic receptors have been proposed to lead to mossy fiber membrane depolarization. How this phenomenon alters the excitability of synaptic boutons, the shape of presynaptic action potentials, Ca2+ influx and neurotransmitter release has remained elusive, but high-resolution live imaging of individual varicosities and direct patch-clamp recordings have begun to shed light on these phenomena. Presynaptic GABAA and kainate receptors have also been reported to facilitate the induction of long-term potentiation at mossy fiber—CA3 synapses. Although mossy fibers are highly specialized, some of the principles emerging at this connection may apply elsewhere in the CNS.

Endogenous zinc depresses GABAergic transmission via T-type Ca2+ channels and broadens the time window for integration of glutamatergic inputs in dentate granule cells

•  Zinc inhibits ionotropic receptors commonly found at central synapses, as well as a wide variety of voltage‐activated ion channels that modulate neuronal excitability and neurotransmitter release. •  We found that zinc chelation facilitated GABAergic signalling in dentate granule cells and that blocking T‐type Ca2+ channel activity abolished this effect. Zinc chelation reduced spike threshold, increased spike width and shifted the input–output relationship in dentate interneurones, which is consistent with increased excitability. •  In granule cells, zinc chelation narrowed the window for the integration of glutamatergic inputs originating from perforant path synapses. •  These results demonstrate that zinc modulates dentate interneurone function and regulates spike routing to local and hippocampal targets.