Prenatal protein malnutrition results in increased frequency of miniature inhibitory synaptic currents in rat CA1 pyramidal cells

There is growing evidence for an effect of prenatal protein malnutrition on the GABAergic neurotransmitter system in the rat hippocampus and associated structures. In the present study, we examined the functional electrophysiological consequences of observed alterations in GABA(A) and benzodiazepine receptor systems. Whole-cell patch clamp recordings of spontaneous and of miniature inhibitory postsynaptic currents (mIPSCs) generated by CA1 pyramidal cells were performed in in vitro hippocampal slices prepared from control and prenatally protein malnourished adult male rats. The characteristics of spontaneous synaptic currents were unaltered by the prenatal insult, as were the amplitudes and kinetics of GABA(A) receptor-mediated mIPSCs. The frequency of mIPSCs, however, was significantly increased in CA1 pyramidal cells in slices prepared from prenatally malnourished vs. control rats. The effect of the benzodiazepine receptor agonist chlordiazepoxide on the characteristics of mIPSCs was also examined and found to be the same in cells from both nutritional groups. The increased frequency of mIPSCs together with the lack of a change in amplitude, kinetics, or modulation by benzodiazepines of mIPSCs in response to prenatal protein malnutrition indicate a presynaptic locus of effect of this insult.     

Sensitivity of synaptic GABA(A) receptors to allosteric modulators in hippocampal oriens-alveus interneurons

GABA(A) receptors are heteropentamers that are heterogeneously distributed at different synapses in the central nervous system. Although the modulation of GABA(A) receptors received much attention in hippocampal pyramidal cells, information is scarce regarding the pharmacology of these receptors in inhibitory interneurons. We investigated the pharmacological properties of GABA(A)-mediated miniature inhibitory postsynaptic currents (mIPSCs) using whole-cell voltage clamp recordings in two morphologically identified types of hippocampal CA1 interneurons, horizontal and vertical cells of stratum oriens-alveus. The negative modulators zinc (200 microM) and furosemide (600 microM) significantly decreased the amplitude of mIPSCs. Benzodiazepine agonists also produced significant effects: 10 microM zolpidem increased the amplitude, rise time, and decay time constant (decay tau) of mIPSCs, whereas 10 microM flunitrazepam affected similarly the amplitude and decay tau, but not the rise time. The neurosteroid allopregnanolone (10 microM) prolonged the decay tau of mIPSCs. Since these modulators act on different GABA(A) receptor subunits, this pharmacological profile suggests that GABA(A) receptors at spontaneously active inhibitory synapses onto vertical and horizontal interneurons are heterogeneous and formed by co-assembly of different combinations of subunits (alpha(1-5)beta(1-3)gamma(1-3)). Furthermore, these synaptic GABA(A) receptors appear in large part pharmacologically similar to those of pyramidal cells.     

Cell type- and synapse-specific variability in synaptic GABAA receptor occupancy

The degree of postsynaptic type A gamma-aminobutyric acid receptor (GABAA receptor) occupancy was investigated by using the benzodiazepine agonist zolpidem. This drug increases the affinity of GABAA receptors for gamma-aminobutyric acid (GABA) at room temperature (Perrais & Ropert 1999, J. Neurosci., 19, 578) leading to an enhancement of synaptic current amplitudes if receptors are not fully occupied by the released transmitter. We recorded miniature inhibitory postsynaptic currents (mIPSCs) from eight different cell types in three brain regions of rats and mice. Receptors in every cell type were benzodiazepine sensitive, as 10-20 microM zolpidem prolonged the decays of mIPSCs (151-184% of control). The amplitude of the GABAA receptor-mediated events was significantly enhanced in dentate granule cells, CA1 pyramidal cells, hippocampal GABAergic interneurons, cortical layer V pyramidal cells, cortical layer V interneurons, and in cortical layer II/III interneurons. An incomplete postsynaptic GABAA receptor occupancy is thus predicted in these cells. In contrast, zolpidem induced no significant change in mIPSC amplitudes recorded from layer II/III pyramidal cells, suggesting full GABAA receptor occupancy. Moreover, different degrees of receptor occupancy could be found at distinct GABAergic synapses on a given cell. For example, of the two distinct populations of zolpidem-sensitive mIPSCs recorded in olfactory bulb granule cells, the amplitude of only one type was significantly enhanced by the drug. Thus, at synapses that generate mIPSCs, postsynaptic receptor occupancy is cell type and synapse specific, reflecting local differences in the number of receptors or in the transmitter concentration in the synaptic cleft.     

Benzodiazepine tolerance at GABAergic synapses on hippocampal CA1 pyramidal cells

Modulation of GABA function following 1 week oral administration of flurazepam (FZP) was investigated in chloride-loaded, rat hippocampal CA1 pyramidal neurons. Rats were sacrificed 2 or 7 days after ending drug treatment, when anticonvulsant tolerance was present or absent in vivo, respectively. Spontaneous (s)IPSCs and miniature (m)IPSCs were recorded using whole-cell voltage-clamp techniques. s/mIPSCs were bicuculline-sensitive, voltage-dependent, and reversed their polarity at 0 mV, the predicted E(Cl-). Comparisons of s/mIPSCs between FZP-treated and control groups were made at Vh = -90, -70, and -50 mV. The frequency of sIPSCs, but not mIPSCs, was significantly decreased in FZP-treated neurons 2 days, but not 7 days, after FZP treatment, suggesting a decrease in interneuron activity. These conclusions were supported by the negative findings of additional studies of [3H]GABA release from hippocampal slices and [3H]GABA uptake from hippocampal synaptosomes. The lack of change in the paired-pulse depression of GABA(B)-mediated IPSPs suggested that autoreceptor function was also not impaired following chronic FZP treatment. A large reduction in both sIPSC and mIPSC amplitude (60%) in FZP-treated neurons, the absence of mIPSCs in one-third of FZP-treated cells, and a measurable reduction in synaptic and unitary conductance confirmed that postsynaptic GABA(A) receptor function was profoundly impaired in FZP-treated CA1 neurons. Zolpidem, an alpha1-selective benzodiazepine receptor ligand, enhanced mIPSC amplitude and decay, but its ability to prolong mIPSC decay was reduced in FZP-treated neurons. Several pre- and postsynaptic changes at GABAergic synapses on CA1 pyramidal cells might be related to the decreased tonic GABA inhibition in FZP-treated CA1 neurons associated with the expression of benzodiazepine anticonvulsant tolerance.     

Nicotine reverses GABAergic inhibition of long-term potentiation induction in the hippocampal CA1 region

Nicotine is known to enhance cognitive function but the mechanism is unknown. The present study examined the modulatory effect of nicotine on the induction of long-term potentiation (LTP), a synaptic model of learning and memory. A weak tetanic stimulation consisting of 20 pulses at 100 Hz induced stable LTP in the hippocampal CA1. The induction of LTP was completely blocked if the tetanus was delivered in the presence of muscimol (2.5 microM), a gamma-aminobutyric acid (GABA) receptor agonist. This inhibition was sensitive to, and reversed by, not only nicotinic acetylcholine receptor (nAChR) agonists (nicotine and epibatidine), but also the alpha7 nAChR-selective antagonist methyllycaconitine (MLA). Furthermore, ACh-puff activation of alpha7 nAChRs on feedforward interneurons induced inhibitory postsynaptic currents in pyramidal cells that were blocked by nicotine or MLA. In addition, nicotine reduced field monosynaptic inhibitory postsynaptic potentials in the presence of MLA. These results suggest not only two pathways of nicotine-induced disinhibition of pyramidal cells, one involving desensitization of alpha7 nAChRs and the other involving non-alpha7 nAChRs, but also two potential mechanisms underlying the modulatory effect of nicotine on LTP induction, both reducing GABAergic inhibition, thereby indirectly increasing the excitability of pyramidal cells.     

Local circuit synaptic interactions between CA1 pyramidal cells and interneurons in the kainate-lesioned hyperexcitable hippocampus.

Following kainate (KA)-induced lesions of subfield CA3--a lesion relevant to human temporal lobe epilepsy--remaining pyramidal cells in CA1 display synchronous hyperexcitability associated with a loss of synaptic inhibition. Despite this loss, inhibitory interneurons in CA1 remain viable, and the density and function of GABAergic receptors on the CA1 pyramidal cells are maintained at approximately normal levels. To further evaluate inhibition in this system, the authors examined interactions between pyramidal cells and inhibitory interneurons in paired intracellular recordings. Recordings were carried out in rat hippocampal slices 2-4 weeks following bilateral intraventricular KA injections. The frequency of synaptic interactions between CA1 basket cells and pyramidal cells was lower in hyperexcitable slices than in controls; both synapses in the recurrent inhibitory circuit appeared to be involved. No recurrent excitatory interactions were seen between pyramidal cell pairs in lesioned or normal slices. The weakened interconnections between pyramidal cells and interneurons are consistent with the decreased inhibition previously found in this model. Unexpectedly, strong stimulation, which may directly activate local inhibitory circuitry, was effective in reducing hyperexcitability in KA-lesioned slices. These data suggest that development of recurrent excitatory connections among CA1 hippocampal pyramidal cells contribute little to tissue excitability, and support the hypothesis that a functional uncoupling between inhibitory interneurons and CA1 pyramidal cells is responsible for the seizure-like activity typical of KA-lesioned hippocampus. The data are also consistent with the hypothesis that in the KA model, the structural circuitry needed for inhibition in CA1 is maintained, and can be functionally activated by appropriate stimuli.     

Intra-hippocampal tonic inhibition influences formalin pain-induced pyramidal cell suppression, but not excitation in dorsal field CA1 of rat

It has been hypothesized that intra-hippocampal GABAergic inhibitory interneurons mediate formalin pain-induced suppression of dorsal hippocampal CA1 pyramidal cell discharge. The present study performed on anaesthetized rats tested the hypothesis by disrupting GABAergic mechanisms with intra-hippocampal administration of the GABA(A) receptor antagonist bicuculline methiodide, applied either dorsally into the pyramidal cell layer and stratum oriens (dorsal-bicuculline) or ventrally into the region of apical dendrites (ventral-bicuculline). It was found that ventral-, but not dorsal-bicuculline attenuated formalin-induced suppression of pyramidal cell extracellular discharge. The antagonism was selective in such a way that the excitation of pyramidal cell was unaffected. Interestingly, ventral-bicuculline strongly disinhibited CA1 pyramidal cells and shifted the distribution of their spontaneous discharge to values higher than the control group. However, dorsal-bicuculline disinhibited the local CA1 interneurons that were strongly excited on injection of formalin. Overall, the findings favour the notion that tonic GABA(A) receptor mechanisms located in the region of apical dendrites facilitate formalin-induced pyramidal cell suppression by masking the background excitatory drive impinging on the pyramidal cells. Interestingly, both the attenuation of formalin-induced inhibition and facilitation of basal discharge of CA1 pyramidal cells by ventral-bicuculline are similar to the effects seen previously with the destruction of medial septal cholinergic neurons. This convergence of effects strengthens the proposal that the network of medial septal cholinergic neurons and hippocampal GABAergic interneurons influence formalin pain-induced CA1 pyramidal cell suppression. In addition, the data point to a non-overlapping excitatory drive whose strength is unaffected by the inhibitory drive that underpins formalin suppression.     

Early sequential formation of functional GABA(A) and glutamatergic synapses on CA1 interneurons of the rat foetal hippocampus

During postnatal development of CA1 pyramidal neurons, GABAergic synapses are excitatory and established prior to glutamatergic synapses. As interneurons are generated before pyramidal cells, we have tested the hypothesis that the GABAergic interneuronal network is operative before glutamate pyramidal neurons and provides the initial patterns of activity. We patch-clamp recorded interneurons in foetal (69 neurons) and neonatal P0 (162 neurons) hippocampal slices and performed a morphofunctional analysis of biocytin-filled neurons. At P0, three types of interneurons were found: (i) non-innervated "silent" interneurons (5%) with no spontaneous or evoked synaptic currents; (ii) G interneurons (17%) with GABA(A) synapses only; and (iii) GG interneurons with GABA and glutamatergic synapses (78%). Relying on the neuronal capacitance, cell body size and arborization of dendrites and axons, the three types of interneurons correspond to three stages of development with non-innervated neurons and interneurons with GABA(A) and glutamatergic synapses being, respectively, the least and the most developed. Recordings from both pyramidal neurons and interneurons in foetuses (E18-20) revealed that the majority of interneurons (65%) had functional synapses whereas nearly 90% of pyramidal neurons were quiescent. Therefore, interneurons follow the same GABA-glutamate sequence of synapse formation but earlier than the principal cells. Interneurons are the source and the target of the first synapses formed in the hippocampus and are thus in a position to modulate the development of the hippocampus in the foetal stage.     

Reduction of GABAB inhibitory postsynaptic potentials by serotonin via pre- and postsynaptic mechanisms in CA3 pyramidal cells of rat hippocampus in vitro.

The action of serotonin (5-HT) on GABAergic synaptic transmission was investigated with intracellular recordings in CA3 pyramidal cells of rat hippocampal slices. Local application of 5-HT (500 microM) hyperpolarized CA3 pyramidal cells, decreased cellular input resistance, and reduced slow afterhyperpolarizations. Serotonin attenuated the late (GABAB) component of polysynaptic inhibitory postsynaptic potentials (IPSPs; 47% of control) without affecting the early (GABAA) component. During bath application of the excitatory amino acid antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) (20 microM) and 2-amino-5-phosphonovalerate (AP-5) (40 microM), 5-HT similarly decreased the amplitude of the late (GABAB) component (17% of control) of monosynaptic IPSPs but did not affect the early (GABAA) component. The mean reversal potentials of poly- and monosynaptic IPSPs were unaffected by 5-HT. The conductance increases associated with the late component of poly- and monosynaptic IPSPs were reduced by 5-HT. Hyperpolarizing responses evoked in CA3 pyramidal cells by somatic applications of gamma-aminobutyric acid (GABA) were unaffected by 5-HT. During bath application of bicuculline (20-50 microM), hyperpolarizing responses elicited by dendritic GABA application were reduced by 5-HT (71% of control). The effect of 5-HT on these direct GABAB hyperpolarizations (29% decrease in response) does not appear sufficient to fully account for the effect of 5-HT on late GABAB IPSPs (53-83% decrease in response). Therefore, 5-HT may reduce GABAB IPSPs in CA3 pyramidal cells 1) by a postsynaptic action on pyramidal cells and 2) by a selective presynaptic action on GABAergic interneurons mediating the GABAB IPSP. This presynaptic action of 5-HT does not appear to involve excitatory afferents onto inhibitory interneurons.     

Early establishment of multiple release site connectivity between interneurons and pyramidal neurons in the developing hippocampus

The strength of the synaptic transmission between two neurons critically depends on the number of release sites connecting the neurons. Here we examine the development of connectivity between gamma-aminobutyric acid (GABA)ergic interneurons and CA1 pyramidal neurons in the hippocampus. GABAergic postsynaptic currents (PSCs) were recorded in whole-cell voltage-clamped CA1 pyramidal neurons. By comparing spontaneous and miniature (action potential-independent) GABAergic PSCs, we found that multiple release site connectivity is established already at the first postnatal day and that the degree of connectivity remains unaltered into adulthood. During the same time there is a dramatic increase in the number of GABAergic synapses on each pyramidal neuron as indicated by the increase in frequency of miniature GABAergic PSCs. These results indicate that during development a given interneuron contacts an increasing number of target pyramidal neurons but with the same multiple release site connectivity. It has been shown previously that the connectivity between CA3 and CA1 pyramidal neurons is initially restricted to one release site, and develops gradually. The present result thus suggests different mechanisms to govern the maturation of excitatory and inhibitory synaptic transmissions.     

Pre- and postsynaptic effects of norepinephrine on γ-aminobutyric acid-mediated synaptic transmission in layer 2/3 of the rat auditory cortex

Noradrenergic terminals from the locus coeruleus release norepinephrine (NE) throughout most brain areas, including the auditory cortex, where they affect neural processing by modulating numerous cellular properties including the inhibitory γ-aminobutyric acid (GABA)ergic transmission. We recently demonstrated that NE affects GABAergic signaling onto cortical pyramidal cells in a complex manner. In this study, we used a combination of patch-clamp recording and immunohistochemical techniques to identify the synaptic site and the location of the adrenergic receptors involved in the modulation of GABAergic signaling in cortical layer 2/3 of the rat. Our results showed that NE increases the frequency of spike-independent miniature inhibitory postsynaptic currents (mIPSCs), as well as the probability of release of unitary inhibitory postsynaptic currents (IPSCs) obtained with patch-clamp pair-recordings. The pharmacology of mIPSCs and the identification of adrenergic receptors in neurons containing the GABAergic marker parvalbumin (PV) suggest that NE increases the presynaptic probability of GABA release by activating α(2) - and β-receptors on PV-positive neurons. On the contrary, bath-applied NE or phenylephrine, decreased the current mediated by pressure application of the GABA(A) -receptor agonist muscimol, as well as the amplitude-but not the frequency-of mIPSCs, indicating that activation of postsynaptic α(1) adrenoceptors reversibly depressed GABAergic currents. We speculate that while a generalized postsynaptic decrease of GABAergic inhibition might decrease the synaptic activation threshold for pyramidal neurons corresponding to an alert state, NE might promote perception and sensory binding by facilitating lateral inhibition as well as the production of γ-oscillations by a selective enhancement of perisomatic inhibition.     

Action of tachykinins in the rat hippocampus: modulation of inhibitory synaptic transmission

Substance P and other neuropeptides of the tachykinin family can powerfully excite CA1 hippocampal interneurons present in the CA1 region. In the present work we show that, by exciting hippocampal interneurons, tachykinins can indirectly inhibit pyramidal neurons. We found that tachykinins caused a decrease in the inhibitory synaptic current interval and an increase in the inhibitory synaptic current amplitude in almost all pyramidal neurons tested. This effect was tetrodotoxin sensitive. Tachykinins did not alter the frequency or amplitude of miniature inhibitory synaptic currents and were without effect on evoked inhibitory synaptic currents. Thus, these neuropeptides acted at the somatodendritic membrane of GABAergic interneurons, rather than at their axon terminals. The effect of substance P on spontaneous inhibitory synaptic currents could be mimicked by a selective agonist of NK1 receptors, but not by selective agonists of NK2 and NK3 receptors. It was suppressed by an NK1 receptor antagonist. In CA1 interneurons located in stratum radiatum, substance P generated a sustained tetrodotoxin-insensitive inward current or induced membrane depolarization and action potential firing. This direct excitatory action was mediated by NK1 receptors. Current-voltage relationships indicate that the net tachykinin-evoked current reversed in polarity at or near the K+ equilibrium potential, suggesting that a suppression of a resting K+ conductance was involved. By increasing the excitability of CA1 GABAergic interneurons, tachykinins can powerfully facilitate the inhibitory synaptic input to pyramidal neurons. This indirect inhibition could play a role in regulating short-term and/or long-term synaptic plasticity, promoting neuronal circuit synchronization or, in some physiopathological situations, influencing epileptogenesis.     

Enkephalin inhibition of inhibitory input to CA1 and CA3 pyramidal neurons in the hippocampus

Enkephalin-induced excitation in the hippocampus has been attributed to the attenuation of inhibitory input as well as to augmentation of excitatory input to pyramidal neurons. We have further examined these possible mechanisms of enkephalin action, as well as the possibility that enkephalins may be affecting intrinsic membrane properties, by recording intracellularly from CA1 and CA3 pyramidal cells in the guinea pig hippocampal brain slice preparation. It was observed that the inhibitory synaptic potential was significantly decreased in the presence of leucine enkephalin and D-alanine, D-leucine-enkephalin (DADL), whereas the excitatory synaptic potential, revealed by block of the inhibitory postsynaptic potential (IPSP) by bicuculline, was unaltered. In addition, the response of pyramidal cells to pressure-applied GABA was unaffected by enkephalin, as were the voltage-dependent membrane conductances. The increase in excitability which was observed in both field potential and intracellular recordings to drop application of DADL must, then, be due to a purely presynaptic block of inhibitory interneurons in both the CA1 and CA3 areas of the hippocampus.     

Anatomically heterogeneous populations of CB1 cannabinoid receptor‐expressing interneurons in the CA3 region of the hippocampus show homogeneous input–output characteristics

A subpopulation of GABAergic cells in cortical structures expresses CB₁ cannabinoid receptors (CB₁) on their axon terminals. To understand the function of these interneurons in information processing, it is necessary to uncover how they are embedded into neuronal circuits. Therefore, the proportion of GABAergic terminals expressing CB₁ and the morphological and electrophysiological properties of CB₁‐immunoreactive interneurons should be revealed. We investigated the ratio and the origin of CB₁‐expressing inhibitory boutons in the CA3 region of the hippocampus. Using immunocytochemical techniques, we estimated that ～40% of GABAergic axon terminals in different layers of CA3 also expressed CB₁. To identify the inhibitory cell types expressing CB₁ in this region, we recorded and intracellularly labeled interneurons in hippocampal slices. CB₁‐expressing interneurons showed distinct axonal arborization, and were classified as basket cells, mossy‐fiber‐associated cells, dendritic‐layer‐innervating cells or perforant‐path‐associated cells. In each morphological category, a substantial variability in axonal projection was observed. In contrast to the diverse morphology, the active and passive membrane properties were found to be rather similar. Using paired recordings, we found that pyramidal cells displayed large and fast unitary postsynaptic currents in response to activating basket and mossy‐fiber‐associated cells, while they showed slower and smaller synaptic events in pairs originating from interneurons that innervate the dendritic layer, which may be due to dendritic filtering. In addition, CB₁ activation significantly reduced the amplitude of the postsynaptic currents in each cell pair tested. Our data suggest that CB₁‐expressing interneurons with different axonal projections have comparable physiological characteristics, contributing to a similar proportion of GABAergic inputs along the somato‐dendritic axis of CA3 pyramidal cells. © 2014 Wiley Periodicals, Inc.     

Effect of transient cerebral ischemia on γ-aminobutyric acidA receptor α1-subunit–immunoreactive interneurons in the gerbil CA1 hippocampus

Following transient cerebral ischemia, pyramidal cells within area CA1 of the hippocampus exhibit delayed neuronal death. While interneurons within this sector continue to survive long-term, there is evidence that some interneurons in area CA1 are vulnerable to damage. To determine the nature of vulnerability in a neurochemically heterogeneous population of interneurons throughout area CA1, we examined the labeling of γ-aminobutyric acid (GABA)ergic interneurons with an antibody to the GABA_A receptor α₁-subunit 1–35 days following cerebral ischemia in the Mongolian gerbil. Unlike some other GABA interneuron markers, this antibody labels both the dendrites and soma of interneurons, allowing dendritic structure to be examined. Three to four days following ischemia, the pyramidal cells in area CA1 had degenerated, and the α₁-subunit–positive interneurons in all layers of area CA1 had developed severely beaded dendrites. At longer survival times (21–35 days), the α₁-subunit–immunolabeled dendrites of these interneurons had a fragmented appearance. In contrast, interneurons bordering str. oriens and alveus typically exhibited normal dendritic morphology. Despite the pathologic changes, there was no evidence of interneuron loss in area CA1 up to 35 days post-ischemia. Normal interneuron morphology was also observed in area CA3 and dentate gyrus, regions where neither pyramidal neurons nor granule cells, respectively, die following 5 min of cerebral ischemia. To determine if the ischemia-induced changes in interneuron morphology could be prevented, diazepam was administered 30 and 90 min following ischemia. Diazepam produces long-term neuroprotection of area CA1 pyramidal neurons. In gerbils sacrificed 35 days after ischemia, diazepam markedly attenuated the dendritic beading of the area CA1 interneurons. In addition, the dendrites did not display the fragmented labeling by the α₁-subunit antibody. Thus, despite their long-term survival, CA1 hippocampal interneurons in the gerbil can express severe structural abnormalities after transient cerebral ischemia coincident with pyramidal cell degeneration, and the injury to the dendrites can be prevented by the neuroprotectant diazepam. Hippocampus 1997; 7:511–523. © 1997 Wiley-Liss, Inc.     

Impaired firing and sodium channel function in CA1 hippocampal interneurons after transient cerebral ischemia.

Although interneurons in area CA1 of the hippocampus are less vulnerable to cerebral ischemia than CA1 pyramidal cells, it is not clear whether their relatively intact cellular morphology implies preservation of normal function. As maintenance of cellular excitability and firing properties is essential for interneurons to regulate neural networks, we investigated these aspects of interneuronal function after transient cerebral ischemia in rats. Cerebral ischemia in rats was induced for 8 mins by a combination of bilateral common carotid artery occlusion and hypovolemic hypotension, and whole cell patch clamp recordings were made in hippocampal slices prepared 24 h after reperfusion. Interneurons located within stratum pyramidale of area CA1 exhibited normal membrane properties and action potentials under these conditions. However, their excitability had declined, as evidenced by an increased action potential threshold and a rightward shift in the relationship between injected depolarizing current and firing rate. Voltage-clamp experiments revealed that transient cerebral ischemia reduced the peak Na(+) current and shifted Na(+) channel activation to more depolarized values, but did not alter steady-state inactivation of the channel. Double immunofluorescence cytochemistry showed that transient cerebral ischemia also reduced Na(v)1.1 subunit immunoreactivity in interneurons that coexpressed parvalbumin. We conclude that transient cerebral ischemia renders CA1 interneurons less excitable, that depressed excitability involves impaired Na(+) channel activation and that Na(+) channel dysfunction is explained, at least in part, by reduced expression of the Na(v)1.1 subunit. These changes may promote interneuron survival, but might also contribute to pyramidal cell death.     

Selective depression of GABA-mediated IPSPs by somatostatin in area CA1 of rabbit hippocampal slices

In area CA1 of hippocampus, a subpopulation of gamma-aminobutyric acid (GABA)-containing interneurons that make synaptic contacts on pyramidal cells also contains the neuropeptide, somatostatin. The effects of GABA and somatostatin on hippocampal pyramidal cells have been investigated separately, but it is not known whether an interaction occurs between these co-localized substances. We demonstrate that somatostatin has a potent inhibitory effect on GABA-mediated synaptic potentials which hyperpolarize pyramidal cells. This effect may be relevant to the well-documented epileptogenicity of the hippocampus, as well as the phenomenon of long-term potentiation, which is a well-studied example of synaptic plasticity.     

Loss of calbindin-immunoreactivity in CA1 hippocampal stratum radiatum and stratum lacunosum-moleculare interneurons in the aged rat

Alterations in hippocampal circuitry may underly age-related learning and memory impairment. We showed in a previous study that the GABA_B-mediated slow inhibitory postsynaptic potential (IPSP) induced in CA1 pyramidal neurons by electrical stimulation of stratum radiatum, is depressed in the hippocampus of the aged rat. This could be due to alterations in GABAergic interneuron functions. We report in this study that the number of hippocampal calbindin-immunoreactive (CaBP-IR) GABAergic interneurons is decreased in the aged rat. The mean number of CaBP-IR interneurons per slice decreases by 50% in the aged rat. The most severe loss was observed in the stratum radiatum of CA1 (78%), with a less consistent loss of immunoreactivity in CA3 (35%). In contrast, the mean number of interneurons containing parvalbumin (PV), was not significantly decreased in the aged rat. Our results show a loss of CaBP immunoreactivity in a population of GABAergic interneurons, which might be related to an altered function of these interneurons and consequently of GABAergic synaptic transmission in the aged rat. In contrast, PV immunoreactivity in interneurons located close to the pyramidal layer does not decrease in the hippocampus of the aged rat.     

Tonically active NMDA receptors – a signalling mechanism critical for interneuronal excitability in the CA1 stratum radiatum

In contrast to tonic extrasynaptic γ‐aminobutyric acid (GABA)_A receptor‐mediated signalling, the physiological significance of tonic extrasynaptic N‐methyl‐d ‐aspartate (NMDA) receptor (NMDAR)‐mediated signalling remains uncertain. In this study, reversible open‐channel blockers of NMDARs, memantine and phencyclidine (PCP) were used as tools to examine tonic NMDAR‐mediated signalling in rat hippocampal slices. Memantine in concentrations up to 10 μm had no effect on synaptically evoked NMDAR‐mediated responses in pyramidal neurons or GABAergic interneurons. On the other hand, 10 μm memantine reduced tonic NMDAR‐mediated currents in GABAergic interneurons by approximately 50%. These tonic NMDAR‐mediated currents in interneurons contributed significantly to the excitability of the interneurons as 10 μm memantine reduced the disynaptic inhibitory postsynaptic current in pyramidal cells by about 50%. Moreover, 10 μm memantine, but also PCP in concentrations ≤ 1 μm , increased the magnitude of the population spike, likely because of disinhibition. The relatively higher impact of tonic NMDAR‐mediated signalling in interneurons was at least partly explained by the expression of GluN2D‐containing NMDARs, which was not observed in mature pyramidal cells. The current results are consistent with the idea that low doses of readily reversible NMDAR open‐channel blockers preferentially inhibit tonically active extrasynaptic NMDARs, and they suggest that tonically active NMDARs contribute more prominently to the intrinsic excitation in GABAergic interneurons than in pyramidal cells. It is proposed that this specific difference between interneurons and pyramidal cells can explain the disinhibition caused by the Alzheimer's disease medication memantine.     

GABAergic control of synaptic summation in hippocampal CA1 pyramidal neurons.

The primary function of neurons is to integrate synaptic inputs and to transmit the results to other cells. It was shown previously that separate excitatory inputs to hippocampal pyramidal neurons are summated nonlinearly. In the hippocampus, responses of pyramidal neurons are influenced by GABAergic inputs in feed-forward or feedback manner, and also by oscillatory network activities. It is likely that these GABAergic inputs regulate the way synaptic inputs are summated. To examine the roles of GABAergic inputs on synaptic summation, we made whole-cell recordings from the cell bodies of CA1 pyramidal neurons in rat hippocampal slices while stimulating two independent input pathways with short interstimulus intervals, and examined the manner by which postsynaptic potentials were summated. We found that: 1) the summation of the perforant pathway and the Schaffer collateral pathway inputs was sublinear when the interval between two inputs was shorter than 30 ms, 2) the blockade of GABA(A) receptors partially suppressed the sublinearity, and 3) further blockade of GABA(B) receptors removed the sublinearity totally. We also found that 4) the summation was superlinear under the concomitant blockade of GABA(A) and GABA(B) receptors when the two inputs arrived with no delay. Thus our study demonstrates that GABAergic inputs are responsible for keeping the summation of two separate inputs on CA1 pyramidal neurons sublinear.