The Role of AMPARs in the Maturation and Integration of Caudal Ganglionic Eminence-Derived Interneurons into Developing Hippocampal Microcircuits.

In the hippocampal CA1, caudal ganglionic eminence (CGE)-derived interneurons are recruited by activation of glutamatergic synapses comprising GluA2-containing calcium-impermeable AMPARs and exert inhibitory regulation of the local microcircuit. However, the role played by AMPARs in maturation of the developing circuit is unknown. We demonstrate that elimination of the GluA2 subunit (GluA2 KO) of AMPARs in CGE-derived interneurons, reduces spontaneous EPSC frequency coupled to a reduction in dendritic glutamatergic synapse density. Removal of GluA1&2&3 subunits (GluA1-3 KO) in CGE-derived interneurons, almost completely eliminated sEPSCs without further reducing synapse density, but increased dendritic branching. Moreover, in GluA1-3 KOs, the number of interneurons invading the hippocampus increased in the early postnatal period but converged with WT numbers later due to increased apoptosis. However, the CCK-containing subgroup increased in number, whereas the VIP-containing subgroup decreased. Both feedforward and feedback inhibitory input onto pyramidal neurons was decreased in GluA1-3 KO. These combined anatomical, synaptic and circuit alterations, were accompanied with a wide range of behavioural abnormalities in GluA1-3 KO mice compared to GluA2 KO and WT. Thus, AMPAR subunits differentially contribute to numerous aspects of the development and maturation of CGE-derived interneurons and hippocampal circuitry that are essential for normal behaviour.

Afferent specific role of NMDA receptors for the circuit integration of hippocampal neurogliaform cells.

Appropriate integration of GABAergic interneurons into nascent cortical circuits is critical for ensuring normal information processing within the brain. Network and cognitive deficits associated with neurological disorders, such as schizophrenia, that result from NMDA receptor-hypofunction have been mainly attributed to dysfunction of parvalbumin-expressing interneurons that paradoxically express low levels of synaptic NMDA receptors. Here, we reveal that throughout postnatal development, thalamic, and entorhinal cortical inputs onto hippocampal neurogliaform cells are characterized by a large NMDA receptor-mediated component. This NMDA receptor-signaling is prerequisite for developmental programs ultimately responsible for the appropriate long-range AMPAR-mediated recruitment of neurogliaform cells. In contrast, AMPAR-mediated input at local Schaffer-collateral synapses on neurogliaform cells remains normal following NMDA receptor-ablation. These afferent specific deficits potentially impact neurogliaform cell mediated inhibition within the hippocampus and our findings reveal circuit loci implicating this relatively understudied interneuron subtype in the etiology of neurodevelopmental disorders characterized by NMDA receptor-hypofunction.Proper brain function depends on the correct assembly of excitatory and inhibitory neurons into neural circuits. Here the authors show that during early postnatal development in mice, NMDAR signaling via activity of long-range synaptic inputs onto neurogliaform cells is required for their appropriate integration into the hippocampal circuitry.

Persistent inhibitory circuit defects and disrupted social behaviour following in utero exogenous cannabinoid exposure.

Placental transfer of Δ9-tetrahydrocannabinol (THC) during pregnancy has the potential to interfere with endogenous cannabinoid (CB) regulation of fetal nervous system development in utero. Here we examined the effect of maternal CB intake on mouse hippocampal interneurons largely focusing on cholecystokinin-expressing interneurons (CCK-INTs), a prominent CB subtype-1 receptor (CB1R) expressing neuronal population throughout development. Maternal treatment with THC or the synthetic CB1R agonist WIN55,212-2 (WIN) produced a significant loss of CCK-INTs in the offspring. Further, residual CCK-INTs in animals prenatally treated with WIN displayed decreased dendritic complexity. Consistent with these anatomical deficits, pups born to CB-treated dams exhibited compromised CCK-INT-mediated feedforward and feedback inhibition. Moreover, pups exposed to WIN in utero lacked constitutive CB1R-mediated suppression of inhibition from residual CCK-INTs and displayed altered social behavior. Our findings add to a growing list of potential cell/circuit underpinnings that may underlie cognitive impairments in offspring of mothers that abuse marijuana during pregnancy.

Burst firing induces postsynaptic LTD at developing mossy fibre-CA3 pyramid synapses.

Synaptic development is an activity-dependent process utilizing coordinated network activity to drive synaptogenesis and subsequent refinement of immature connections. Hippocampal CA3 pyramidal neurons (PYRs) exhibit intense burst firing (BF) early in development, concomitant with the period of mossy fibre (MF) development. However, whether developing MF-PYR synapses utilize PYR BF to promote MF synapse maturation remains unknown. Recently, we demonstrated that transient tonic depolarization of postsynaptic PYRs induces a persistent postsynaptic form of long-term depression (depolarization-induced long-term depression, DiLTD) at immature MF-PYR synapses. DiLTD induction is NMDAR independent but does require postsynaptic Ca(2+) influx through L-type voltage gated Ca(2+) channels (L-VGCCs), and is expressed as a reduction in AMPAR function through the loss of GluR2-lacking AMPARs present at immature MF-PYR synapses. Here we examined whether more physiologically relevant phasic L-VGCC activation by PYR action potential (AP) BF activity patterns can trigger DiLTD. Using combined electrophysiological and Ca(2+) imaging approaches we demonstrate that PYR BF effectively drives L-VGCC activation and that brief periods of repetitive PYR BF, produced by direct current injection or intrinsic network activity induces NMDAR-independent LTD by promoting Ca(2+) influx through the activated L-VGCCs. This BF induced LTD, just like DiLTD, is specific for developing MF-PYR synapses, is PICK1 dependent, and is expressed postsynaptically. Our results demonstrate that DiLTD can be induced by phasic L-VGCC activation driven by PYR BF, suggesting the engagement of natural PYR network activity patterns for MF synapse maturation.

Regulation of Kv1 subunit expression in oligodendrocyte progenitor cells and their role in G1/S phase progression of the cell cycle.

Proliferative oligodendrocyte progenitor cells (OPs) express large, delayed outward-rectifying K(+) currents (I(K)), whereas nondividing immature and mature oligodendrocytes display much smaller I(K). Here, we show that up-regulation of I(K) occurs in G(1) phase of the cell cycle in purified cultured OPs and is the result of an RNA synthesis-dependent, selective increase of the K(+) channel subunit proteins Kv1.3 and Kv1.5. In oligodendrocyte cells acutely isolated from developing rat brain, a decrease of cyclin D expression is observed as these cells mature along their lineage. This is accompanied by a decrease in Kv1.3 and Kv1.5 subunit expression, suggesting a role for these subunits in the proliferative potential of OPs in situ. I(K) expressed in OPs in subventricular zone and developing white matter in acutely isolated slice preparations were selectively blocked by antagonists of Kv1.3, illustrating the functional presence of this subunit in situ. Interestingly, Kv1.3 block inhibited S-phase entry of both purified OPs in culture and in tissue slice cultures. Thus, we employ both in vitro and in situ experimental approaches to show that (i) RNA-dependent synthesis of Kv1.3 and Kv1.5 subunit proteins occurs in G(1) phase of the OP cell cycle and is responsible for the observed increase in I(K), and (ii) currents through Kv1.3-containing channels play a crucial role in G(1)/S transition of proliferating OPs.

Differential synaptic processing separates stationary from transient inputs to the auditory cortex.

Sound features are blended together en route to the central nervous system before being discriminated for further processing by the cortical synaptic network. The mechanisms underlying this synaptic processing, however, are largely unexplored. Intracortical processing of the auditory signal was investigated by simultaneously recording from pairs of connected principal neurons in layer II/III in slices from A1 auditory cortex. Physiological patterns of stimulation in the presynaptic cell revealed two populations of postsynaptic events that differed in mean amplitude, failure rate, kinetics and short-term plasticity. In contrast, transmission between layer II/III pyramidal neurons in barrel cortex were uniformly of large amplitude and high success (release) probability (Pr). These unique features of auditory cortical transmission may provide two distinct mechanisms for discerning and separating transient from stationary features of the auditory signal at an early stage of cortical processing.

Kv3 channels: voltage-gated K+ channels designed for high-frequency repetitive firing.

Analysis of the Kv3 subfamily of K(+) channel subunits has lead to the discovery of a new class of neuronal voltage-gated K(+) channels characterized by positively shifted voltage dependencies and very fast deactivation rates. These properties are adaptations that allow these channels to produce currents that can specifically enable fast repolarization of action potentials without compromising spike initiation or height. The short spike duration and the rapid deactivation of the Kv3 currents after spike repolarization maximize the quick recovery of resting conditions after an action potential. Several neurons in the mammalian CNS have incorporated into their repertoire of voltage-dependent conductances a relatively large number of Kv3 channels to enable repetitive firing at high frequencies - an ability that crucially depends on the special properties of Kv3 channels and their impact on excitability.

Interneurons unbound.

Local-circuit, gamma-aminobutyric acid-releasing inhibitory interneurons of the hippocampus and cortex have traditionally been considered as the regulators of principal neuron activity--the yin to the excitatory yang. Recent evidence indicates that, in addition to that role, their network connectivity and the properties of their intrinsic voltage-gated currents are finely tuned to permit inhibitory interneurons to generate and control the rhythmic output of large populations of both principal cells and other populations of inhibitory interneurons. This review brings together recently described properties and emerging principles of interneuron function that indicate a much more complex role for these cells than just providers of inhibition.

Differential mechanisms of transmission at three types of mossy fiber synapse.

The axons of the dentate gyrus granule cells, the so-called mossy fibers, innervate their inhibitory interneuron and pyramidal neuron targets via both anatomically and functionally specialized synapses. Mossy fiber synapses onto inhibitory interneurons were comprised of either calcium-permeable (CP) or calcium-impermeable (CI) AMPA receptors, whereas only calcium-impermeable AMPA receptors existed at CA3 principal neuron synapses. In response to brief trains of high-frequency stimuli (20 Hz), pyramidal neuron synapses invariably demonstrated short-term facilitation, whereas interneuron EPSCs demonstrated either short-term facilitation or depression. Facilitation at all CI AMPA synapses was voltage independent, whereas EPSCs at CP AMPA synapses showed greater facilitation at -20 than at -80 mV, consistent with a role for the postsynaptic unblock of polyamines. At pyramidal cell synapses, mossy fiber EPSCs possessed marked frequency-dependent facilitation (commencing at stimulation frequencies >0.1 Hz), whereas EPSCs at either type of interneuron synapse showed only moderate frequency-dependent facilitation or underwent depression. Presynaptic metabotropic glutamate receptors (mGluRs) decreased transmission at all three synapse types in a frequency-dependent manner. However, after block of presynaptic mGluRs, transmission at interneuron synapses still did not match the dynamic range of EPSCs at pyramidal neuron synapses. High-frequency stimulation of mossy fibers induced long-term potentiation (LTP), long-term depression (LTD), or no change at pyramidal neuron synapses, interneuron CP AMPA synapses, and CI AMPA synapses, respectively. Induction of LTP or LTD altered the short-term plasticity of transmission onto both pyramidal cells and interneuron CP AMPA synapses by a mechanism consistent with changes in release probability. These data reveal differential mechanisms of transmission at three classes of mossy fiber synapse made onto distinct targets.

Snap-25 is polarized to axons and abundant along the axolemma: an immunogold study of intact neurons.

SNAP-25, synaptosomal associated protein of 25 kDa, is reported to be a t-SNARE (target receptor associated with the presynaptic plasma membrane) involved in the docking and fusion of synaptic vesicles. We present here the first ultrastructural localization of SNAP-25 in intact neurons by pre-embedding EM immunocytochemistry in rat brains, hippocampal slice cultures, and PC12 cells. In differentiated neurons, SNAP-25 labeling was clearly membrane-associated. The labeling was most prominent in the plasma membrane of axons and excluded from the plasma membranes of soma and dendrites. Furthermore, SNAP-25 did not appear to be restricted to the synaptic junctions. SNAP-25 labeling was seen in the cytoplasm of the soma and large dendrites, mostly associated with the Golgi complexes. There were also some SNAP-25 labeled tubulo-vesicular structures in the cytoplasm of the soma and the axons, but rarely in the smaller dendrites. In PC12 cells, after 5-10 minutes of high potassium (75 mM) stimulation in the presence of HRP, SNAP-25 labeling appeared, additionally, on HRP-filled early endosomes. After a longer (20-30 minutes) HRP incubation, most of the later stage endosomes and lysosomes were loaded with HRP but they were negative for SNAP-25. These results suggest that SNAP-25 is sorted out of these late endosomal compartments, and that the bulk of the SNAP-25 protein is probably recycled back to the axolemma from the early endosomes. In contrast, in those samples which were incubated with HRP for longer periods, there were still some SNAP-25-positive vesicular structures which were HRP-negative. These structures most likely represent anterograde vesicles that carry newly synthesized SNAP-25 from the soma to the axolemma by axonal transport. SNAP-25 appears to be sorted at the Golgi complex to reach the axolemma specifically. Its widespread distribution all along the axolemma does not support the view of SNAP-25 as a t-SNARE limited for synaptic exocytosis.

H2 histamine receptor-phosphorylation of Kv3.2 modulates interneuron fast spiking.

Histamine-containing neurons of the tuberomammilary nucleus project to the hippocampal formation to innervate H1 and H2 receptors on both principal and inhibitory interneurons. Here we show that H2 receptor activation negatively modulates outward currents through Kv3.2-containing potassium channels by a mechanism involving PKA phosphorylation in inhibitory interneurons. PKA phosphorylation of Kv3.2 lowered the maximum firing frequency of inhibitory neurons, which in turn negatively modulated high-frequency population oscillations recorded in principal cell layers. All these effects were absent in a Kv3.2 knockout mouse. These data reveal a novel pathway for histamine-dependent regulation of high-frequency oscillations within the hippocampal formation.

Target-specific expression of pre- and postsynaptic mechanisms.

Target-specific expression of pre- and postsynaptic mechanisms of synaptic transmission has been shown in a variety of central neurons by a number of laboratories. These data have demonstrated that synaptic transmission between single axons diverging onto distinct target neurons can behave independently, differentially influencing activity in the target neuron. Similarly, single neurons are capable of manufacturing molecularly distinct ligand-gated receptors and targeting them to synapses innervated by distinct converging afferent projections. A picture is emerging consistent with a role for both pre- and postsynaptic mechanisms in influencing the target-specific nature of transmission at numerous diverse synapses throughout the mammalian CNS. This target specificity adds another level of complexity in unravelling the roles played by individual neurons within a computational network. To begin to understand the coordinated activity of large ensembles of neurons it is becoming clear that the nature of transmission between individual pre- and postsynaptic elements within a circuit must first be understood for each and every neural element involved.

Multiple forms of feedback inhibition by str. oriens inhibitory interneurons?

In the CA1 region of the hippocampus pyramidal neuron basilar dendrites extend into the stratum oriens-alveus while the apical dendrites project deep into the stratum lacunosum-moleculare, a distance several hundred micrometres in extent. This extended dendritic arbor provides a large surface area for afferent input. For example, the axons of CA3 pyramidal neurons synapse onto pyramidal cell dendritic spines across approximately 2/3 of the apical dendritic tree. In contrast a diverse population of local-circuit GABAergic inhibitory interneurons selectively innervate specific postsynaptic domains of principal cells (for review see Freund & Buzsaki, 1996). These cells target their axons either to the axon initial segment, somata, or proximal and distal dendrites, with each cell type implicated in a particular operational role. However, rigid classification of the numerous subpopulations has been problematic (see Parra et al. 1998 for further discussion).

Hippocampal abnormalities and enhanced excitability in a murine model of human lissencephaly.

Human cortical heterotopia and neuronal migration disorders result in epilepsy; however, the precise mechanisms remain elusive. Here we demonstrate severe neuronal dysplasia and heterotopia throughout the granule cell and pyramidal cell layers of mice containing a heterozygous deletion of Lis1, a mouse model of human 17p13.3-linked lissencephaly. Birth-dating analysis using bromodeoxyuridine revealed that neurons in Lis1+/- murine hippocampus are born at the appropriate time but fail in migration to form a defined cell layer. Heterotopic pyramidal neurons in Lis1+/- mice were stunted and possessed fewer dendritic branches, whereas dentate granule cells were hypertrophic and formed spiny basilar dendrites from which the principal axon emerged. Both somatostatin- and parvalbumin-containing inhibitory neurons were heterotopic and displaced into both stratum radiatum and stratum lacunosum-moleculare. Mechanisms of synaptic transmission were severely disrupted, revealing hyperexcitability at Schaffer collateral-CA1 synapses and depression of mossy fiber-CA3 transmission. In addition, the dynamic range of frequency-dependent facilitation of Lis1+/- mossy fiber transmission was less than that of wild type. Consequently, Lis1+/- hippocampi are prone to interictal electrographic seizure activity in an elevated [K(+)](o) model of epilepsy. In Lis1+/- hippocampus, intense interictal bursting was observed on elevation of extracellular potassium to 6.5 mM, a condition that resulted in only minimal bursting in wild type. These anatomical and physiological hippocampal defects may provide a neuronal basis for seizures associated with lissencephaly.

Frequency-dependent regulation of rat hippocampal somato-dendritic excitability by the K+ channel subunit Kv2.1.

The voltage-dependent potassium channel subunit Kv2.1 is widely expressed throughout the mammalian CNS and is clustered primarily on the somata and proximal dendrites, but not axons, of both principal neurones and inhibitory interneurones of the cortex and hippocampus. This expression pattern suggests that Kv2.1-containing channels may play a role in the regulation of pyramidal neurone excitability. To test this hypothesis and to determine the functional role of Kv2. 1-containing channels, cultured hippocampal slices were incubated with antisense oligonucleotides directed against Kv2.1 mRNA. Western blot analysis demonstrated that Kv2.1 protein content of cultured slices decreased > 90 % following 2 weeks of treatment with antisense oligonucleotides, when compared with either control missense-treated or untreated cultures. Similarly, Kv2.1 immunostaining was selectively decreased in antisense-treated cultures. Sustained outward potassium currents, recorded in both whole-cell and outside-out patch configurations, demonstrated a selective reduction of amplitude only in antisense-treated CA1 pyramidal neurones. Under current-clamp conditions, action potential durations were identical in antisense-treated, control missense-treated and untreated slices when initiated by low frequency stimulation (0.2 Hz). In contrast, spike repolarization was progressively prolonged during higher frequencies of stimulation (1 Hz) only in cells from antisense-treated slices. Similarly, action potentials recorded during electrographic interictal activity in the 'high [K+]o' model of epilepsy demonstrated pronounced broadening of their late phase only in cells from antisense-treated slices. Consistent with the frequency-dependent spike broadening, calcium imaging experiments from single CA1 pyramidal neurones revealed that high frequency Schaffer collateral stimulation resulted in a prolonged elevation of dendritic [Ca2+]i transients only in antisense-treated neurones. These studies demonstrate that channels containing Kv2.1 play a role in regulating pyramidal neurone somato-dendritic excitability primarily during episodes of high frequency synaptic transmission.

Voltage-activated K+ channels and membrane depolarization regulate accumulation of the cyclin-dependent kinase inhibitors p27(Kip1) and p21(CIP1) in glial progenitor cells.

Neural cell development is regulated by membrane ion channel activity. We have previously demonstrated that cell membrane depolarization with veratridine or blockage of K+ channels with tetraethylammonium (TEA) inhibit oligodendrocyte progenitor (OP) proliferation and differentiation (); however the molecular events involved are largely unknown. Here we show that forskolin (FSK) and its derivative dideoxyforskolin (DFSK) block K+ channels in OPs and inhibit cell proliferation. The antiproliferative effects of TEA, FSK, DFSK, and veratridine were attributable to OP cell cycle arrest in G1 phase. In fact, (1) cyclin D accumulation in synchronized OP cells was not affected by K+ channel blockers or veratridine; (2) these agents prevented OP cell proliferation only if present during G1 phase; and (3) G1 blockers, such as rapamycin and deferoxamine, mimicked the anti-proliferative effects of K+ channel blockers. DFSK also prevented OP differentiation, whereas FSK had no effect. Blockage of K+ channels and membrane depolarization also caused accumulation of the cyclin-dependent kinase inhibitors p27(Kip1) and p21(CIP1) in OP cells. The antiproliferative effects of K+ channel blockers and veratridine were still present in OP cells isolated from INK4a-/- mice, lacking the cyclin-dependent kinase inhibitors p16(INK4a) and p19(ARF). Our results demonstrate that blockage of K+ channels and cell depolarization induce G1 arrest in the OP cell cycle through a mechanism that may involve p27(Kip1) and p21(CIP1) and further support the conclusion that OP cell cycle arrest and differentiation are two uncoupled events.

Glutamatergic synapses onto hippocampal interneurons: precision timing without lasting plasticity.

In the hippocampal formation GABAergic inhibitory interneurons have 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 innervation from glutamatergic neurons and it has been much debated whether these synapses show mechanisms of long-term plasticity similar to those found at principal-cell synapses. Recent findings support the lack of conventional forms of LTP and LTD in most interneurons, partly owing to the distinct anatomical and neurochemical features of interneuronal excitatory synapses. The uncommon properties of excitatory synapses on interneurons might be required for their functioning as accurate and reliable neuronal oscillators.

Neurotransmitter receptor activation triggers p27(Kip1 )and p21(CIP1) accumulation and G1 cell cycle arrest in oligodendrocyte progenitors.

We examined the pathways that link neurotransmitter receptor activation and cell cycle arrest in oligodendrocyte progenitors. We had previously demonstrated that glutamate receptor activation inhibits oligodendrocyte progenitor proliferation and lineage progression. Here, using purified oligodendrocyte progenitors and cerebellar slice cultures, we show that norepinephrine and the beta-adrenergic receptor agonist isoproterenol also inhibited the proliferation, but in contrast to glutamate, isoproterenol stimulated progenitor lineage progression, as determined by O4 and O1 antibody staining. This antiproliferative effect was specifically attributable to a beta-adrenoceptor-mediated increase in cyclic adenosine monophosphate, since analogs of this cyclic nucleotide mimicked the effects of isoproterenol on oligodendrocyte progenitor proliferation, while alpha-adrenoceptor agonists were ineffective. Despite the opposite effects on lineage progression, both isoproterenol and the glutamate receptor agonist kainate caused accumulation of the cyclin-dependent kinase inhibitors p27(Kip1)and p21(CIP1), and G1 arrest. Studies with oligodendrocyte progenitor cells from INK4a-/- mice indicated that the G1 cyclin kinase inhibitor p16(INK4a) as well as p19(ARF)were not required for agonist-stimulated proliferation arrest. Our results demonstrate that beta-adrenergic and glutamatergic receptor activation inhibit oligodendrocyte progenitor proliferation through a mechanism that may involve p27(Kip1) and p21(CIP1); but while neurotransmitter-induced accumulation of p27(Kip1) is associated with cell cycle arrest, it does not by itself promote oligodendrocyte progenitor differentiation.

Graded reduction of Pafah1b1 (Lis1) activity results in neuronal migration defects and early embryonic lethality.

Heterozygous mutation or deletion of the beta subunit of platelet-activating factor acetylhydrolase (PAFAH1B1, also known as LIS1) in humans is associated with type I lissencephaly, a severe developmental brain disorder thought to result from abnormal neuronal migration. To further understand the function of PAFAH1B1, we produced three different mutant alleles in mouse Pafah1b1. Homozygous null mice die early in embryogenesis soon after implantation. Mice with one inactive allele display cortical, hippocampal and olfactory bulb disorganization resulting from delayed neuronal migration by a cell-autonomous neuronal pathway. Mice with further reduction of Pafah1b1 activity display more severe brain disorganization as well as cerebellar defects. Our results demonstrate an essential, dosage-sensitive neuronal-specific role for Pafah1b1 in neuronal migration throughout the brain, and an essential role in early embryonic development. The phenotypes observed are distinct from those of other mouse mutants with neuronal migration defects, suggesting that Pafah1b1 participates in a novel pathway for neuronal migration.