Reduced tonic inhibition in the dentate gyrus contributes to chronic stress‐induced impairments in learning and memory

It is well established that stress impacts the underlying processes of learning and memory. The effects of stress on memory are thought to involve, at least in part, effects on the hippocampus, which is particularly vulnerable to stress. Chronic stress induces hippocampal alterations, including but not limited to dendritic atrophy and decreased neurogenesis, which are thought to contribute to chronic stress‐induced hippocampal dysfunction and deficits in learning and memory. Changes in synaptic transmission, including changes in GABAergic inhibition, have been documented following chronic stress. Recently, our laboratory demonstrated shifts in E_GABA in CA1 pyramidal neurons following chronic stress, compromising GABAergic transmission and increasing excitability of these neurons. Interestingly, here we demonstrate that these alterations are unique to CA1 pyramidal neurons, since we do not observe shifts in E_GABA following chronic stress in dentate gyrus granule cells. Following chronic stress, there is a decrease in the expression of the GABA_A receptor (GABA_AR) δ subunit and tonic GABAergic inhibition in dentate gyrus granule cells, whereas there is an increase in the phasic component of GABAergic inhibition, evident by an increase in the peak amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs). Given the numerous changes observed in the hippocampus following stress, it is difficult to pinpoint the pertinent contributing pathophysiological factors. Here we directly assess the impact of a reduction in tonic GABAergic inhibition of dentate gyrus granule cells on learning and memory using a mouse model with a decrease in GABA_AR δ subunit expression specifically in dentate gyrus granule cells (Gabrd/Pomc mice). Reduced GABA_AR δ subunit expression and function in dentate gyrus granule cells is sufficient to induce deficits in learning and memory. Collectively, these findings suggest that the reduction in GABA_AR δ subunit‐mediated tonic inhibition in dentate gyrus granule cells contributes, at least in part, to deficits in learning and memory associated with chronic stress. These findings have significant implications regarding the pathophysiological mechanisms underlying impairments in learning and memory associated with stress and suggest a role for GABA_AR δ subunit containing receptors in dentate gyrus granule cells. © 2016 Wiley Periodicals, Inc.     

Hippocampal GABAergic dysfunction in a rat chronic mild stress model of depression

In major depression, one line of research indicates that a dysfunctional GABAergic inhibitory system is linked to the appearance of depressive symptoms. However, as the mechanistic details of such GABAergic deficit are largely unknown, we undertook a functional investigation of the GABAergic system in the rat chronic mild stress model of depression. Adult rats were exposed to an eight‐week long stress protocol leading to anhedonic‐like behavior. In hippocampal brain slices, phasic, and tonic GABA_A receptor‐mediated currents in dentate gyrus granule cells were examined using patch‐clamp recordings. In granule cells, the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was reduced to 41% in anhedonic‐like rats, which was associated with a reduced probability of evoked GABA release. Using immunohistochemical analysis, there was no change in the number of parvalbumin‐positive interneurons in the dentate gyrus. Notably, we observed a 60% increase in THIP‐activated tonic GABA_A mediated current in anhedonic‐like rats, suggesting an upregulation of extrasynaptic GABA_A receptors. Finally, five weeks treatment with the antidepressant escitalopram partially reversed the sIPSCs frequency. In summary, we have revealed a hippocampal dysfunction in the GABAergic system in the chronic mild stress model of depression in rats, caused by a reduction in action potential‐dependent GABA release. Since the function of the GABAergic system was improved by antidepressant treatment, in parallel with behavioral read outs, it suggests a role of the GABAergic system in the pathophysiology of depression. © 2010 Wiley‐Liss, Inc.     

Tonic GABA(A) receptor-mediated currents in human brain

GABA(A) receptors can mediate both phasic (synaptic) and tonic (extrasynaptic) forms of inhibition. It has been proposed that tonic inhibition plays a critical part in controlling neuronal and network excitability. Although tonic GABA(A) receptor-mediated currents have been well characterized in rodents, their existence in human tissue has yet to be demonstrated. Here we show that tonic currents can be recorded from human tissue obtained from patients undergoing temporal lobectomies. Tonic GABA(A) receptor-mediated currents were present in pyramidal cells and interneurons in layer V-VI of temporal neocortex and granule cells in the dentate gyrus. These tonic currents have cell type-specific pharmacologies, opening up the possibility of targeted therapeutics.     

Granule cell disinhibition in dentate gyrus of genetically seizure susceptible El mice

Paired-pulse inhibition was investigated electrophysiologically in the dentate gyrus using hippocampal slices from epileptic El mice. At short interpulse intervals (IPIs), the inhibition was 30% in the El, and 90% in the control ddY mice at the ages of 10 and 15 weeks. No difference in inhibition was observed at the age of 5 weeks. Bicuculline, a GABA_A receptor antagonist, attenuated the inhibition during short IPIs in the ddY mice, while in the El mice, phenobarbital and flunitrazepam, which enhance GABA_A receptor function, restored the inhibitory activity comparable to that of the ddY. The disinhibition progressed with growth, closely correlating with seizure development in El mice. These results suggest that decrease in the GABAergic inhibition occurs in the dentate gyrus of the El mice with growth. GABA concentration in the hippocampus was also quantified using HPLC. In El mice, GABA level was significantly lower than that in ddY mice at the ages of 5 and 15 weeks. Thus, the disinhibition observed in the El dentate gyrus at 15 weeks of age does not appear to be directly related to the content of GABA. GABAergic disinhibition suggests possible loss of unknown inhibition control factor(s) in the El dentate gyrus as growth progresses. The growth-dependent disinhibition in the granule cells may be a prerequisite for epileptogenesis in El mice.     

Presynaptic But Not Postsynaptic GABA Signaling at Unitary Mossy Fiber Synapses

Dentate gyrus granule cells have been suggested to corelease GABA and glutamate both in juvenile animals and under pathological conditions in adults. Although mossy fiber terminals (MFTs) are known to express glutamic acid decarboxylase (GAD) in early postnatal development, the functional role of GABA synthesis in MFTs remains controversial, and direct evidence for synaptic GABA release from MFTs is missing. Here, using GAD67-GFP transgenic mice, we show that GAD67 is expressed only in a population of immature granule cells in juvenile animals. We demonstrate that GABA can be released from these cells and modulate mossy fiber excitability through activation of GABAB autoreceptors. However, unitary postsynaptic currents generated by individual, GAD67-expressing granule cells are purely glutamatergic in all postsynaptic cell types tested. Thus GAD67 expression does not endow dentate gyrus granule cells with a full GABAergic phenotype and GABA primarily instructs the pre- rather than the postsynaptic element.     

Spatiotemporal specificity of GABAA receptor-mediated regulation of adult hippocampal neurogenesis

GABAergic transmission regulates adult neurogenesis by exerting negative feedback on cell proliferation and enabling dendrite formation and outgrowth. Further, GABAergic synapses target differentiating dentate gyrus granule cells prior to formation of glutamatergic connections. GABA(A) receptors (GABA(A) Rs) mediating tonic (extrasynaptic) and phasic (synaptic) transmission are molecularly and functionally distinct, but their specific role in regulating adult neurogenesis is unknown. Using global and single-cell targeted gene deletion of subunits contributing to the assembly of GABA(A) Rs mediating tonic (α4, δ) or phasic (α2) GABAergic transmission, we demonstrate here in the dentate gyrus of adult mice that GABA(A) Rs containing α4, but not δ, subunits mediate GABAergic effects on cell proliferation, initial migration and early dendritic development. In contrast, α2-GABA(A) Rs cell-autonomously signal to control positioning of newborn neurons and regulate late maturation of their dendritic tree. In particular, we observed pruning of distal dendrites in immature granule cells lacking the α2 subunit. This alteration could be prevented by pharmacological inhibition of thrombospondin signaling with chronic gabapentin treatment, shown previously to reduce glutamatergic synaptogenesis. These observations point to homeostatic regulation of inhibitory and excitatory inputs onto newborn granule cells under the control of α2-GABA(A) Rs. Taken together, the availability of distinct GABA(A) R subtypes provides a molecular mechanism endowing spatiotemporal specificity to GABAergic control of neuronal maturation in adult brain.     

The development of GABAergic neurons in the rat hippocampal formation. An immunocytochemical study

Recent studies have indicated that hippocampal GABAergic neurons in both the dentate gyrus and Ammon's horn are generated prenatally. Although the adult distribution of GABAergic neurons has been previously described by numerous investigators, the early postnatal appearance of these neurons has not been described. In the present study, immunocytochemical methods were used to localize GABAergic neurons with antisera to both GABA and its synthesizing enzyme, glutamate decarboxylase (GAD). The GABA-positive neurons appeared at the earliest postnatal day (PND) examined, 4 PND. In contrast, GAD-positive cells were not observed until 6 PND, and the number of these neurons remained less than that of the GABA-positive neurons until 14 PND. These findings indicated that immunocytochemically detectable amounts of GAD were not present in many young GABAergic neurons. Both GABA- and GAD-positive hippocampal neurons showed two large increases in number during the 4-8 PND and 12-16 PND time periods, and they reached about 90% of adult levels before 18 PND. The regional distribution of GABA- and GAD-positive neurons throughout the hippocampal formation was homogeneous for all ages examined except 4 PND. At this age, the GABA-positive cells appeared in clusters in the proximal CA3 and the distal CA1 relative to the dentate gyrus. In addition, the number of hippocampal neurons immunostained in adult preparations for both antisera to GABA and GAD showed a similar number and distribution. The data on the developmental appearance of GABA and GAD immunoreactivities are consistent with biochemical data for the development of GABA concentration and GAD activity in the hippocampal formation. Together, these data provide important information about the functional maturation of the hippocampal GABAergic system in the first 3 weeks of rat brain development.     

Similar GABAergic inputs in dentate granule cells born during embryonic and adult neurogenesis

Neurogenesis in the dentate gyrus of the hippocampus follows a unique temporal pattern that begins during embryonic development, peaks during the early postnatal stages and persists through adult life. We have recently shown that dentate granule cells born in early postnatal and adult mice acquire a remarkably similar afferent connectivity and firing behavior, suggesting that they constitute a homogeneous functional population [Laplagne et al. (2006)PLoS Biol., 4, e409]. Here we extend our previous study by comparing mature neurons born in the embryonic and adult hippocampus, with a focus on intrinsic membrane properties and gamma-aminobutyric acid (GABA)ergic synaptic inputs. For this purpose, dividing neuroblasts of the ventricular wall were retrovirally labeled with green fluorescent protein at embryonic day 15 (E15), and progenitor cells of the subgranular zone were labeled with red fluorescent protein in the same mice at postnatal day 42 (P42, adulthood). Electrophysiological properties of mature neurons born at either stage were then compared in the same brain slices. Evoked and spontaneous GABAergic postsynaptic responses of perisomatic and dendritic origin displayed similar characteristics in both neuronal populations. Miniature GABAergic inputs also showed similar functional properties and pharmacological profile. A comparative analysis of the present data with our previous observations rendered no significant differences among GABAergic inputs recorded from neurons born in the embryonic, early postnatal and adult mice. Yet, embryo-born neurons showed a reduced membrane excitability, suggesting a lower engagement in network activity. Our results demonstrate that granule cells of different age, location and degree of excitability receive GABAergic inputs of equivalent functional characteristics.     

GABA is depolarizing in hippocampal dentate granule cells of the adolescent and adult rats

GABAergic signaling in hippocampal pyramidal neurons undergoes a switch from depolarizing to hyperpolarizing during early neuronal development. Whether such a transformation of GABAergic action occurs in dentate granule cells (DGCs), located at the first stage of the hippocampal trisynaptic circuit, is unclear. Here, we use noninvasive extracellular recording to monitor the effect of synaptically released GABA on the DGC population. We find that GABAergic responses in adolescent and adult rat DGCs are still depolarizing from rest. Using a morphologically realistic DGC model, we show that GABAergic action, depending on its precise timing and location, can have either an excitatory or inhibitory role in signal processing in the dentate gyrus.     

Robust tonic GABA currents can inhibit cell firing in mouse newborn neocortical pyramidal cells

Within the hippocampus and neocortex, GABA is considered to be excitatory in early development due to a relatively depolarized Cl^? reversal potential (E_Cl). Although the depolarizing nature of synaptic GABAergic events has been well established, it is unknown whether cortical tonic currents mediated by extrasynaptically located GABA_A receptors (GABA_ARs) are also excitatory. Here we examined the development of tonic currents in the neocortex and their effect on neuronal excitability. Mean tonic current, recorded from layer 5 (L5) pyramidal cells of the mouse somatosensory cortex, is robust in newborns [postnatal day (P)2–4] then decreases dramatically by the second postnatal week (P7–10 and P30–40). Pharmacological studies, in combination with Western blot analysis, show that neonatal tonic currents are partially mediated by the GABA_AR α5 subunit, and probably the δ subunit. In newborns, the charge due to tonic current accounts for nearly 100% of the total GABA charge, a contribution that decreases to < 50% in mature tissue. Current clamp recordings show that tonic current contributes to large fluctuations in the membrane potential that may disrupt its stability. Bath application of 5 μM GABA, to induce tonic currents, markedly decreased cell firing frequency in most recorded cells while increasing it in others. Gramicidin perforated patch recordings show heterogeneity in E_Cl recorded from P2–5 L5 pyramidal cells. Together, these findings demonstrate that tonic currents activated by low GABA concentrations can dominate GABAergic transmission in newborn neocortical pyramidal cells and that tonic currents can exert heterogeneous effects on neuronal excitability.     

Stress and hippocampal neurogenesis.

The dentate gyrus of the hippocampal formation develops during an extended period that begins during gestation and continues well into the postnatal period. Furthermore, the dentate gyrus undergoes continual structural remodeling in adulthood. The production of new granule neurons in adulthood has been documented in a number of mammalian species, ranging from rodents to primates. The late development of this brain region makes the dentate gyrus particularly sensitive to environmental and experience-dependent structural changes. Studies have demonstrated that the proliferation of granule cell precursors, and ultimately the production of new granule cells, are dependent on the levels of circulating adrenal steroids. Adrenal steroids inhibit cell proliferation in the dentate gyrus during the early postnatal period and in adulthood. The suppressive action of glucocorticoids on cell proliferation is not direct but occurs through an NMDA receptor-dependent excitatory pathway. Stressful experiences, which are known to elevate circulating levels of glucocorticoids and stimulate hippocampal glutamate release, inhibit the proliferation of granule cell precursors. Chronic stress results in persistent inhibition of granule cell production and changes in the structure of the dentate gyrus, raising the possibility that stress alters hippocampal function through this mechanism. This review considers the unusual developmental profile of the dentate gyrus and its vulnerability to environmental perturbations. The long-term impact of developmental events on hippocampal function is considered.     

Phasic GABAA -receptor activation is required to suppress epileptiform activity in the CA3 region of the immature rat hippocampus

Purpose: Despite the consistent observation that γ‐aminobutyric acid A (GABA_A) receptors mediate excitatory responses at perinatal stages, the role of the GABAergic system in the generation of neonatal epileptiform activity remains controversial. Therefore, we analyzed whether tonic and phasic GABAergic transmission had differential effects on neuronal excitability during early development. Methods: We performed whole cell patch‐clamp and field potential recordings in the CA3 region of hippocampal slices from immature (postnatal day 4–7) rats to analyze the effect of specific antagonists and modulators of tonic and phasic GABAergic components on neuronal excitability. Key Findings: The GABAergic antagonists gabazine (3 μm ) and picrotoxin (100 μm ) induced epileptiform discharges, whereas activation of GABA_A receptors attenuated epileptiform discharges. Under low‐Mg²⁺ conditions, 100 nm gabazine and 1 μm picrotoxin were sufficient to provoke epileptiform activity in 63.2% (n = 19) and 53.8% (n = 26) of the slices, respectively. Whole‐cell patch‐clamp experiments revealed that these concentrations significantly reduced the amplitude of phasic GABAergic postsynaptic currents but had no effect on tonic currents. In contrast, 1‐μm 4,5,6,7‐tetrahydroisoxaz‐olo[5,4‐c]‐pyridin‐3‐ol (THIP) induced a tonic current of ?12 ± 2.5 pA (n = 6) and provoked epileptiform discharges in 57% (n = 21) of the slices. Significance: We conclude from these results that in the early postnatal rat hippocampus a constant phasic synaptic activity is required to control excitability and prevent epileptiform activity, whereas tonic GABAergic currents can mediate excitatory responses. Pharmacologic intervention at comparable human developmental stages should consider these ambivalent GABAergic actions.     

Interactions of neurotransmitter systems during postnatal development of the rat hippocampal formation: effects of cisplatin

The distribution of neuroimmunohistochemical markers for the serotoninergic, noradrenergic, glutamatergic and GABAergic systems (respectively, 5HT(2A)R, β1AR, GluR 2/3 and GAD65/67) was determined in the hippocampal formation at stages PD11, PD17 and PD30 of postnatal development of untreated rats and cisplatin-treated rats after a single injection of the drug at 10days of life. In the different time points the neurons of the dentate gyrus and Cornu Ammonis progressively acquire mature morphological characteristics, and cell genesis, migration of interneurons and differentiation of mossy cells occur. Cisplatin induced decrease in immunoreactivity for most of the selected neurotransmitter markers, thereby altering the postnatal development of circuits in the hippocampal formation. Cisplatin also brought out clear evidence for an interaction between excitatory and inhibitory neurotransmitter markers during the postnatal maturation of cells and fiber projections containing GluR2/3 and GAD65, despite the fact that glutamatergic neurons and GABAergic interneurons are divergent in their source of genesis and in their mode of migration. In fact, GluR2/3 immunofluorescence was increased in the principal cells early, at PD11, possibly to reduce the calcium influx into the cell. Moreover, cisplatin might cause a loss of GABAergic interneurons early and reduction of fiber projections to hippocampal layers due to altered cell migration or to cell injury; late changes, particularly in GAD67 cell number in the dentate gyrus did not result in redistribution or recovery in treated rats. With the use of cisplatin it has been demonstrated here for the first time that the critical differentiation of dentate gyrus hilar β1AR and GluR2/3 mossy cells takes place between PD11 and PD17. Changes in neurotransmitter marker immunopositivity occurred subsequently to cytoarchitectural changes in the dentate gyrus and Cornu Ammonis which were already evident one day after cisplatin injection, suggesting that degeneration and cell loss may have occurred. Cisplatin was found to be a useful tool for following CNS development and for understanding how hippocampal neuronal networks react to injury. Furthermore, cisplatin-induced neurotoxicity may be used to reveal useful information on the genesis, migration and distribution, and differentiation of distinct types of hippocampal neurons.     

Synchronized spike waves in immature dentate gyrus networks

At early developmental stages, immature neuronal networks of the neocortex and hippocampus spontaneously exhibit synchronously oscillating activities, which are believed to play roles in normal circuit maturation. The tissue development of the dentate gyrus (DG) in the hippocampal formation is exceptionally late compared with other brain regions and persists until postnatal periods. Using patch-clamp recording and functional multineuron calcium imaging, we found that the DG networks of postnatal day (P)3-7 mice spontaneously generated traveling waves of action potentials, which were initiated at the upper blade of the granule cell layer and propagated to the lower blade. The propagation was dependent on glutamatergic and electrical synapses, but not on GABAergic receptor activity. Remarkably, the DG waves were almost completely abolished in offspring born to female rats exposed to restraint stress during pregnancy. In the prenatally stressed offspring, DG granule cell dendrites developed normally until P3 and showed atrophy by P9. Thus, the DG waves may be required for the maturation of DG granule cells.     

Decrease in tonic inhibition contributes to increase in dentate semilunar granule cell excitability after brain injury

Brain injury is an etiological factor for temporal lobe epilepsy and can lead to memory and cognitive impairments. A recently characterized excitatory neuronal class in the dentate molecular layer, semilunar granule cell (SGC), has been proposed to regulate dentate network activity patterns and working memory formation. Although SGCs, like granule cells, project to CA3, their typical sustained firing and associational axon collaterals suggest that they are functionally distinct from granule cells. We find that brain injury results in an enhancement of SGC excitability associated with an increase in input resistance 1 week after trauma. In addition to prolonging miniature and spontaneous IPSC interevent intervals, brain injury significantly reduces the amplitude of tonic GABA currents in SGCs. The postinjury decrease in SGC tonic GABA currents is in direct contrast to the increase observed in granule cells after trauma. Although our observation that SGCs express Prox1 indicates a shared lineage with granule cells, data from control rats show that SGC tonic GABA currents are larger and sIPSC interevent intervals shorter than in granule cells, demonstrating inherent differences in inhibition between these cell types. GABA(A) receptor antagonists selectively augmented SGC input resistance in controls but not in head-injured rats. Moreover, post-traumatic differences in SGC firing were abolished in GABA(A) receptor blockers. Our data show that cell-type-specific post-traumatic decreases in tonic GABA currents boost SGC excitability after brain injury. Hyperexcitable SGCs could augment dentate throughput to CA3 and contribute substantively to the enhanced risk for epilepsy and memory dysfunction after traumatic brain injury.     

Evidence for changing positions of GABA neurons in the developing rat dentate gyrus

In recent studies, we demonstrated a distinct change in the distribution of glutamate decarboxylase 67 (GAD67) mRNA-containing neurons within the rat dentate gyrus from embryonic day 20 (E20) to postnatal day 15 (PN15) (Dupuy and Houser, J Comp Neurol 1997;389:402-418). We also observed a similar changing pattern for cells with birthdates of many of the mature GAD-containing neurons in the dentate gyrus (Dupuy and Houser, J Comp Neurol 1997;389:402-418). These observations suggested that some early-appearing GABA neurons within the developing molecular layer of the dentate gyrus may gradually alter their positions to become the mature GABAergic cells along the inner border of the granule cell layer. The goal of the present study was to provide additional evidence for our hypothesis by demonstrating the spatial relationships between GAD-containing neurons and granule cells at progressively older ages during development. In this study, immunohistochemical or in situ hybridization methods for the localization of GAD67 or its mRNA were combined with bromodeoxyuridine birthdating techniques that labeled early-generated granule cells with birthdates on E17. At E20, GAD67-containing neurons were located above the granule cell layer that contained E17 birthdated granule cells. During the first two postnatal weeks, both GAD67 mRNA-containing neurons and early-born granule cells were primarily concentrated within the granule cell layer. Double-labeled neurons were rarely observed, and this suggests that these two groups are separate populations. By PN15-PN30, most GAD67 mRNA-containing neurons were distributed along the base of the granule cell layer, significantly below the E17 birthdated granule cells. These findings support our new hypothesis that mature GABA neurons along the inner border of the granule cell layer reach their positions by migrating or translocating through the developing granule cell layer.     

Partial inactivation of GABAA receptors containing the α5 subunit affects the development of adult‐born dentate gyrus granule cells

Alterations of neuronal activity due to changes in GABA_A receptors (GABA_AR) mediating tonic inhibition influence different hippocampal functions. Gabra5‐null mice and α5 subunit^(H105R) knock‐in mice exhibit signs of hippocampal dysfunction, but are capable of improved performance in several learning and memory tasks. Accordingly, alleviating abnormal GABAergic tonic inhibition in the hippocampal formation by selective α5‐GABA_AR modulators represents a possible therapeutic approach for several intellectual deficit disorders. Adult neurogenesis in the dentate gyrus is an important facet of hippocampal plasticity; it is regulated by tonic GABAergic transmission, as shown by deficits in proliferation, migration and dendritic development of adult‐born neurons in Gabra4‐null mice. Here, we investigated the contribution of α5‐GABA_ARs to granule cell development, using retroviral vectors expressing eGFP for labeling precursor cells in the subgranular zone. Global α5‐GABA_AR knockout (α5‐KO) mice showed no alterations in migration and morphological development of eGFP‐positive granule cells. However, upregulation of α1 subunit‐immunoreactivity was observed in the hippocampal formation and cerebral cortex. In contrast, partial gene inactivation in α5‐heterozygous (α5‐het) mice, as well as single‐cell deletion of Gabra5 in newborn granule cells from α5‐floxed mice, caused severe alterations of migration and dendrite development. In α5‐het mice, retrovirally mediated overexpression of Cdk5 resulted in normal migration and dendritic branching, suggesting that Cdk5 cooperates with α5‐GABA_ARs to regulate neuronal development. These results show that minor imbalance of α5‐GABA_AR‐mediated transmission may have major consequences for neuronal plasticity; and call for caution upon chronic therapeutic use of negative allosteric modulators acting at these receptors.     

A comparison of the ontogeny of excitatory and inhibitory neurotransmission in the CA1 region and dentate gyrus of the rat hippocampal formation.

In vivo electrophysiological experiments were used to chart the ontogeny of excitatory and inhibitory neurotransmission in the hippocampal formation of rats. Using standardized protocols, responses in the dentate gyrus were quantified and systematically compared to similar measurements obtained in the CA1 region. Measurements were taken at numerous ages, ranging from postnatal day (PN) 6 to adults (PN 60). Excitation was monitored by two parameters recorded with extracellular electrodes in response to monosynaptic inputs to CA1 pyramidal cells or to dentate gyrus granule cells: maximum population spike (PSmax) amplitudes and maximum population excitatory postsynaptic potential slopes (pEPSP slopemax). Inhibition was assessed by a paired-pulse protocol to measure maximal inhibition (the potency of inhibition at an interpulse interval of 20 ms) and duration of inhibition (the interpulse interval at which paired-pulse inhibition changed to paired-pulse facilitation). Excitatory parameters matured later in the dentate gyrus than in CA1, consistent with the later appearance of granule cells. Until PN 21, pEPSPmax values in the dentate gyrus paralleled those in CA1; thereafter they diverged with far larger values in the dentate gyrus. Inhibitory parameters reached adult values between PN 14 and 18. In both regions paired-pulse responses consisted of three phases: (1) an initial inhibition; (2) a second facilitatory phase; and (3) a later inhibition. The maximal inhibition in the initial phase was comparable in both regions, but lasted longer in the CA1 region. The facilitation in the second phase was greater in the dentate gyrus, and the inhibition in the third phase was greater in the dentate gyrus. Results are discussed in terms of neurogenesis of principal cells and GABAergic cells in the regions of interest.     

The mechanisms of the strong inhibitory modulation of long-term potentiation in the rat dentate gyrus

The hippocampus is essential for the formation of certain types of memory, and synaptic plasticity such as long-term potentiation (LTP) is widely accepted as a cellular basis of hippocampus-dependent memory. Although LTP in both perforant path-dentate gyrus (DG) granule cell and CA3-CA1 pyramidal cell synapses is similarly dependent on activation of postsynaptic N-methyl-D-aspartate receptors, several reports suggest that modulation of LTP by γ-aminobutyric acid (GABA) receptor-mediated inhibitory inputs is stronger in perforant path-DG granule cell synapses. However, little is known about how different the mechanism and physiological relevance of the GABAergic modulation of LTP induction are among different brain regions. We confirmed that the action of GABA(A) receptor antagonists on LTP was more prominent in the DG, and explored the mechanism introducing such difference by examining two types of GABA(A) receptor-mediated inhibition, i.e. synaptic and tonic inhibition. As synaptic inhibition, we compared inhibitory vs. excitatory monosynaptic responses and their summation during an LTP-inducing stimulus, and found that the balance of the summated postsynaptic currents was biased toward inhibition in the DG. As tonic inhibition, or sustained activation of extrasynaptic GABA(A) receptors by ambient GABA, we measured the change in holding currents of the postsynaptic cells induced by GABA(A) receptor antagonists, and found that the tonic inhibition was significantly stronger in the DG. Furthermore, we found that tonic inhibition was associated with LTP modulation. Our results suggest that both the larger tonic inhibition and the larger inhibitory/excitatory summation balance during conditioning are involved in the stronger inhibitory modulation of LTP in the DG.     

GABA mediates the network activity-dependent facilitation of axonal outgrowth from the newborn granule cells in the early postnatal rat hippocampus

Neural network activity regulates the development of hippocampal newborn granule cells (GCs). Excitatory GABAergic input is known to be a key player in this regulation. Although calcium signaling is thought to be a downstream mediator of GABA, GABA-induced calcium signaling in newborn GCs is not well understood. We investigated Ca(2+) signaling and its regulatory role in axon and dendrite outgrowth in newborn GCs identified in the organotypic slice culture of early postnatal rat hippocampus. Here, we report that hippocampal network activity can induce calcium transients (CaTs) in newborn GCs during the first post-mitotic week via GABAergic inputs. The GABA-induced CaTs were mediated mainly by L-type Ca(2+) channels. Furthermore, we found that inhibiting any step in the signaling pathway, network activity → GABA → L-type Ca(2+) channels, selectively suppressed the axonal outgrowth and pruning of newborn GCs, but not dendritic outgrowth. The GABA(A) receptor blocker bicuculline significantly suppressed axonal outgrowth, despite increasing network activity, thus indicating an essential role of GABAergic inputs. Therefore, we conclude that network activity-dependent GABAergic inputs open L-type Ca(2+) channels and promote axonal outgrowth in newborn GC during the first post-mitotic week.