Opposite effects of glucocorticoid receptor activation on hippocampal CA1 dendritic complexity in chronically stressed and handled animals

Remodeling of synaptic networks is believed to contribute to synaptic plasticity and long‐term memory performance, both of which are modulated by chronic stress. We here examined whether chronic stress modulates dendritic complexity of hippocampal CA1 pyramidal cells, under conditions of basal as well as elevated corticosteroid hormone levels. Slices were prepared from naïve, handled or chronically stressed animals and briefly treated with vehicle or corticosterone (100 nM); neurons were visualized with a fluorescent dye injected into individual CA1 pyramidal cells. We observed that 21 days of unpredictable stress did not affect hippocampal CA1 apical or basal dendritic morphology compared with naïve animals when corticosteroid levels were low. Only when slices from stressed animals were also exposed to elevated corticosteroid levels, a significant reduction in apical (but not basal) dendritic length became apparent. Unexpectedly, animals that were handled or 3 weeks showed a reduction in both apical dendritic length and number of apical branch points when compared with naïve animals. Apical dendritic length and number of branch points were restored to levels found in naïve animals several hours after in vitro treatment with 100 nM corticosterone. All effects of acute corticosterone administration could be prevented by the glucocorticoid receptor antagonist RU38486 given during the last 4 days of the stress or handling protocol. We conclude that brief exposure to high concentrations of corticosterone can differently affect apical dendritic structure, depending on the earlier history of the animal, a process that critically depends on involvement of the glucocorticoid receptor. © 2007 Wiley‐Liss, Inc.     

新生大鼠海马CA1神经元突触反应和树突分枝的关系

应用盲法脑片膜片钳记录并结合biocytin细胞内染色方法 ,研究了新生大鼠 (生后 3～ 5d)离体海马脑片CA1锥体神经元突触反应和树突分枝的关系。发现 ,在生后 3～ 5d锥体神经元的形态呈现多形性。 5 2 %的神经元具有分枝很少的树突 ,并且既无自发性也无诱发性的突触后电流 (postsynapticcurrents,PSCs) ;4 8%的神经元具有较为发达的树突分枝且当刺激海马辐射层时 ,可引起突触反应。而且在半最大刺激强度时引起的PSCs的幅值与神经元顶树突的长度及终末分枝数呈正相关。上述研究结果表明 ,CA1锥体神经元的反应性是与顶树突的分枝状况相关的。     

Nimodipine decreases calcium action potentials in rabbit hippocampal CA1 neurons in an age-dependent and concentration-dependent manner

Intracellular recordings were made from rabbit hippocampal CA1 neurons in vitro using slices from aging and young adult rabbits. Calcium action potentials were studied in the presence of 4 μm tetrodotoxin using electrodes filled with 2M CsCl. Increasing concentrations of the dihydropyridine L-type calcium channel antagonist nimodipine were tested on the amplitude and time course of calcium action potentials. The calcium action potential (AP) consisted of two components: an initial fast phase followed by a slower plateau phase. No difference in the peak amplitude of the initial fast phase was observed between age groups. The amplitude and duration of the slower plateau phase of the calcium AP was significantly larger in aging neurons. Switching to a zero Ca²⁺ medium in the presence of 200 μm CdCl₂ completely blocked the calcium AP. Nimodipine decreased the plateau phase of the calcium AP at concentrations as low as 100 nm in aging neurons and 10 μm in young neurons. Switching to higher concentrations of nimodipine did not reveal any substantially increased block of the calcium AP plateau phase. These data suggest that enhanced calcium influx through L-type calcium channels is largely responsible for the enhanced calcium action potentials observed in aging CA1 neurons. The action of nimodipine in reducing the plateau phase of the calcium action potential may underlie the drug's notable ability to improve learning in hippocampally dependent tasks in aging animals.     

Circadian rhythm of redox state regulates membrane excitability in hippocampal CA1 neurons

Behaviors, such as sleeping, foraging, and learning, are controlled by different regions of the rat brain, yet they occur rhythmically over the course of day and night. They are aligned adaptively with the day‐night cycle by an endogenous circadian clock in the suprachiasmatic nucleus (SCN), but local mechanisms of rhythmic control are not established. The SCN expresses a ~24‐hr oscillation in reduction‐oxidation that modulates its own neuronal excitability. Could circadian redox oscillations control neuronal excitability elsewhere in the brain? We focused on the CA1 region of the rat hippocampus, which is known for integrating information as memories and where clock gene expression undergoes a circadian oscillation that is in anti‐phase to the SCN. Evaluating long‐term imaging of endogenous redox couples and biochemical determination of glutathiolation levels, we observed oscillations with a ~24 hr period that is 180° out‐of‐phase to the SCN. Excitability of CA1 pyramidal neurons, primary hippocampal projection neurons, also exhibits a rhythm in resting membrane potential that is circadian time‐dependent and opposite from that of the SCN. The reducing reagent glutathione rapidly and reversibly depolarized the resting membrane potential of CA1 neurons; the magnitude is time‐of‐day‐dependent and, again, opposite from the SCN. These findings extend circadian redox regulation of neuronal excitability from the SCN to the hippocampus. Insights into this system contribute to understanding hippocampal circadian processes, such as learning and memory, seizure susceptibility, and memory loss with aging.     

Relation between responsiveness to neurotransmitters and complexity of epileptiform activity in rat hippocampal CA1 neurons

PURPOSE: Our previous works suggested that sensitivity of neurons with chaotic firing patterns to stimuli is significantly greater than that in neurons with periodic firing patterns, which shows that responsiveness of neurons may depend on the complexity of the firing series. This study was performed to determine the relation between responsiveness of the hippocampal CA1 neurons with epileptiform activity (EA) to neurotransmitters and their complexity of firing series. METHODS: Firing series of CA1 neurons were recorded extracellularly in rat hippocampal slice. Approximate entropy was used to describe the complexity of the interspike interval (ISI) series. EA was induced by local application of penicillin (1,000 IU/ml). The change of firing rate induced by neurotransmitters (glutamate and gamma-aminobutyric acid) was compared with that of the degree of complexity of ISI series in the process of EA. RESULTS: The excitatory responses to glutamate and the inhibitory responses to gamma-aminobutyric acid in CA1 neurons appeared to be decreased during the process of penicillin-induced EA. However, during this same process, the approximate entropy of the ISI series also was decreased significantly. CONCLUSIONS: The results suggest that the reduced responses to neurotransmitters of the CA1 neurons appear to be closely related to the onset of EA. Furthermore, these neurons show that the changes in responsiveness are closely parallel to the decrease of degree of complexity of firing series during penicillin epileptogenesis.     

Neuropeptide Y reduces orthodromically evoked population spike in rat hippocampal CA1 by a possibly presynaptic mechanism

Application of the brain neuropeptide Y (NPY) to rat hippocampus in vitro reversibly reduced the amplitude of the CA1 population spike evoked by stratum radiatum stimulation. Threshold for the effect was 10(-8) M. NPY had similar effects on single pulse- and paired pulse-evoked population spikes. Antidromic population spikes, evoked from the alveus, were unaffected by NPY. Thus, NPY appears to modulate excitatory transmission in the hippocampus by a presynaptic mechanism.     

海马CA1锥体神经元NMDA及非NMDA受体介导的兴奋性突触后电流的生后变化

应用盲法脑片膜片钳记录方法,研究了幼年大鼠（生后6～21d）及成年大鼠（生后56～70d）离体海马脑片CA1锥体神经元N-甲基-D-门冬氨酸（NMDA）及非NMDA受体介导的兴奋性突触后电流（EPSCs）的生后发育变化。为阻断γ-氨基丁酸（GABA）能抑制性突触活动,灌注液中常规应用GABAA受体拮抗剂荷包牡丹碱（50umol/L）。局部刺激海马辐射层（0.05～0.1Hz）可引起EPSCs。结果显示NMDA受体拮抗剂AP5可减小EPSCs的幅值,减小的程度以幼年大鼠为显著。进一步灌注a-氨基-3羧基-5-甲基异恶唑-4丙酸（AMPA）受体拮抗剂CNQX（20umol/L）可完全阻断残余EPSCs。分析给予AP5前、后EPSCs幅值的大小,可得到NMDA及非NMDA受体介导EPSCs,显示非NMDA受体介导的EPSCs随着发育明显增加,而NMDA万分则降低。上述研究结果表明海马CA1神经元的兴奋性突触活动是由NMDA及非NMDA受体介导的,并且在生后一周内以NMDA成分为主,因此在发育的早期NMDA受体可能更多参与对发育的调节作用。     

Perforant pathway-evoked long-term potentiation of CA1 neurons in the hippocampal slice preparation

Previously, we have presented electrophysiological evidence reaffirming the existence of a controversial hippocampal pathway. These fibers are part of the perforant pathway and terminate directly on the CA1 cells. We now report that, in the hippocampal slice preparation, tetanic stimulation of the perforant pathway produces long-term potentiation (LTP) of CA1 cell responses. LTP of population spikes varied from 150% to 500%. The results were of interest because these axons synapse at distal sites on the apical dendrite. This location is usually thought to be a difficult site to evoke action potentials.     

Orexin‐A increases the firing activity of hippocampal CA1 neurons through orexin‐1 receptors

Orexins including two peptides, orexin‐A and orexin‐B, are produced in the posterior lateral hypothalamus. Much evidence has indicated that central orexinergic systems play numerous functions including energy metabolism, feeding behavior, sleep/wakefulness, and neuroendocrine and sympathetic activation. Morphological studies have shown that the hippocampal CA1 regions receive orexinergic innervation originating from the hypothalamus. Positive orexin‐1 (OX₁) receptors are detected in the CA1 regions. Previous behavioral studies have shown that microinjection of OX₁ receptor antagonist into the hippocampus impairs acquisition and consolidation of spatial memory. However, up to now, little has been known about the direct electrophysiological effects of orexin‐A on hippocampal CA1 neurons. Employing multibarrel single‐unit extracellular recordings, the present study showed that micropressure administration of orexin‐A significantly increased the spontaneous firing rate from 2.96 ± 0.85 to 8.45 ± 1.86 Hz (P < 0.001) in 15 out of the 23 hippocampal CA1 neurons in male rats. Furthermore, application of the specific OX₁ receptor antagonist SB‐334867 alone significantly decreased the firing rate from 4.02 ± 1.08 to 2.11 ± 0.58 Hz in 7 out of the 17 neurons (P < 0.05), suggesting that endogenous orexinergic systems modulate the firing activity of CA1 neurons. Coapplication of SB‐334867 completely blocked orexin‐A–induced excitation of hippocampal CA1 neurons. The PLC pathway may be involved in activation of OX₁ receptor–induced excitation of CA1 neurons. Taken together, the present study's results suggest that orexin‐A produces excitatory effects on hippocampal neurons via OX₁ receptors. © 2016 Wiley Periodicals, Inc.     

Blockade of glucocorticoid receptors rapidly restores hippocampal CA1 synaptic plasticity after exposure to chronic stress

Prolonged exposure to stressful events has been reported to inhibit the ability of hippocampal synapses to increase their synaptic efficacy. Here we tested if these effects could be prevented by blocking activation of glucocorticoid receptors during the last 4 days of the stress paradigm. In order to address this question, animals were exposed to 21 days of variable and inescapable stressors. Handled animals served as controls. During the last 4 days of the stress regime, animals were treated with the glucocorticoid receptor antagonist RU486. We found that 1 day after the last stressor, synaptic plasticity in the CA1 area of hippocampal slices is impaired in chronically stressed animals. Importantly, treating chronically stressed animals with RU486 for 4 days completely prevented this decrease in synaptic potentiation; RU486 treatment of handled controls did not affect potentiation. Treating hippocampal slices from control animals with high levels of corticosterone also impaired synaptic plasticity; this effect was similar for untreated and RU486-treated animals. Treating slices from chronically stressed animals with corticosterone did not further decrease synaptic plasticity. These data indicate that 4 days blockade of the glucocorticoid receptor, during a stress regime, is sufficient to fully restore synaptic plasticity.     

Nicotine exerts differential effects on different CA1 hippocampal cell types

The effects of (−) nicotine hydrogen tartrate (NHT) were examined on several cell types in the CA1 region of rat hippocampus. The results indicate that nicotine may have a preferential net inhibitory effect on basket cells and an excitatory effect on oriens/alveus interneurons. The resultant effects of nicotine on pyramidal cells may thus be a product of complex local circuit interactions.     

Transient hippocampal CA1 lesions in humans impair pattern separation performance

Day‐to‐day life involves the perception of events that resemble one another. For the sufficient encoding and retrieval of similar information, the hippocampus provides two essential computational processes. Pattern separation refers to the differentiation of overlapping memory representations, whereas pattern completion reactivates memories based on noisy or degraded input. Evidence from human and rodent studies suggest that pattern separation specifically relies on neuronal ensemble activity in hippocampal subnetworks in the dentate gyrus and CA3. Although a role for CA1 in pattern separation has been shown in animal models, its contribution in the human hippocampus remains elusive. In order to elucidate the contribution of CA1 neurons to pattern separation, we examined 14 patients with an acute transient global amnesia (TGA), a rare self‐limiting dysfunction of the hippocampal system showing specific lesions to CA1. Patients' pattern separation performance was tested during the acute amnestic phase and follow‐up using an established mnemonic similarity test. Patients in the acute phase showed a profound deficit in pattern separation (p < .05) as well as recognition memory (p < .001) that recovered during follow‐up. Specifically, patients tested in a later stage of the amnesia were less impaired in pattern separation than in recognition memory. Considering the time dependency of lesion‐associated hippocampal deficits in early and late acute stages of the TGA, we showed that the pattern separation function recovered significantly earlier than recognition memory. Our results provide causal evidence that hippocampal CA1 neurons are critical to pattern separation performance in humans.     

Differential developmental refinement of the intrinsic electrophysiological properties of CA1 pyramidal neurons from the rat dorsal and ventral hippocampus

The dorsal and ventral regions of the rat longitudinal hippocampal axis are functionally distinct. That is, each region is associated with different behavioral tasks and disease susceptibilities due to underlying anatomical, and physiological differences. These differences are especially pronounced in area CA1, where significant differences in morphology, synaptic physiology, intrinsic excitability, and gene expression have been reported between CA1 pyramidal neurons from the dorsal (DHC) and ventral hippocampus (VHC). However, despite a significant amount of recent attention, a cogent picture of the intrinsic electrophysiological profile of DHC and VHC neurons has remained elusive, due, in part, to experiments performed on rats at different developmental time points. Moreover, the resulting intrinsic electrophysiological profiles are sufficiently different as to warrant a thorough investigation of the spatial and temporal changes in the intrinsic excitability of CA1 pyramidal neurons across developmental time. Accordingly, in this study, I have characterized the intrinsic electrophysiological properties of CA1 pyramidal neurons from acute hippocampal slices prepared from the DHC and VHC throughout an approximately 3‐week developmental period (P14–P37). DHC and VHC neurons exhibited distinct intra‐region changes (DHC or VHC) and inter‐region differences (DHC versus VHC) in their intrinsic electrophysiological properties, which yielded two developmental timelines: (a) a common developmental timeline describing changes observed in both DHC and VHC neurons, and (b) a differential developmental timeline highlighting unique features observed in DHC neurons. Specifically, DHC neurons exhibited significant inter‐region differences in RMP, input resistance, threshold, and spike frequency adaptation relative to VHC neurons, as well as an intra‐region change in the rebound slope (a proxy for I_h). These observations both integrate and reconcile previous work performed with rats at different developmental stages and suggest a distinct developmental trajectory for DHC neurons that might shed light on the normal physiological functions and disease susceptibility of the DHC.     

Early ischemia enhances action potential-dependent, spontaneous glutamatergic responses in CA1 neurons

Two types of quantal spontaneous neurotransmitter release are present in the nervous system, namely action potential (AP)-dependent release and AP-independent release. Previous studies have identified and characterized AP-independent release during hypoxia and ischemia. However, the relative contribution of AP-dependent spontaneous release to the overall glutamate released during transient ischemia has not been quantified. Furthermore, the neuronal activity that mediates such release has not been identified. Using acute brain slices, we show that AP-dependent release constitutes approximately one-third of the overall glutamate-mediated excitatory postsynaptic potentials/currents (EPSPs/EPSCs) measured onto hippocampal CA1 pyramidal neurons. However, during transient (2 mins) in vitro hypoxia–hypoglycemia, large-amplitude, AP-dependent spontaneous release is significantly enhanced and contributes to 74% of the overall glutamatergic responses. This increased AP-dependent release is due to hyper-excitability in the presynaptic CA3 neurons, which is mediated by the activity of NMDA receptors. Spontaneous glutamate release during ischemia can lead to excitotoxicity and perturbation of neural network functions.     

Pathway specificity of dendritic spine morphology in identified synapses onto rat hippocampal CA1 neurons in organotypic slices

The output of the hippocampus is largely determined by interaction of the three excitatory pathways that impinge on CA1 pyramidal neurons. These synapses, formed by axons of: (1) CA3 pyramidal neurons; (2) neurons of the entorhinal cortex (EC); and (3) neighboring CA1 neurons, are all potentially plastic. Here, we take advantage of the accessibility of the organotypic slice preparation to identify the type of spines with which each of these pathways forms synapses, at different developmental stages. Recent reports have shown that morphology of dendritic spines is activity-dependent with large mushroom spines being thought to represent stronger synaptic connections than thin or stubby spines. Although in a wide range of preparations, mushroom spines represent only 15% of spines across the whole dendritic tree, we find that this proportion is highly pathway specific. Thus in organotypic slices, the axons of CA3 neurons form synapses with mushroom spines on CA1 neurons in approximately 50% of cases, whereas this spine type is rare (<10%) in either of the other two pathways. This high proportion of mushroom spines only occurs after spontaneous excitatory activity in the CA1 cells increases over the second week in vitro. Previous studies suggest that pathway specificity also occurs in vivo. In tissue fixed in vivo, it is the synapses of distal apical dendrites thought to be formed by axons originating in the EC that are richer in mushroom spines. Hence, contrary to previous suggestions, the proportion of mushroom spines is clearly not an intrinsic property of the pathway but rather a characteristic dependent on the environment. We suggest that this is most likely a result of the previous activity of the synapses. The fact that, despite the large differences in pathway specificity between preparations, the overall proportion of different spine types remains unchanged, suggests a strong influence of homeostasis across the network.     

Anoxia-induced depolarization in CA1 hippocampal neurons: role of Na-dependent mechanisms

We have previously shown that (1) removal of extracellular sodium (Na⁺) reduces the anoxia-induced depolarization in neurons in brain-slice preparations and (2) amiloride, which blocks Na⁺-dependent exchangers, prevents anoxic injury in cultured neocortical neurons. Since anoxia-induced depolarization has been linked to neuronal injury, we have examined in this study the role of Na⁺-dependent exchangers and voltage-gated Na⁺ channels in the maintenance of membrane properties of CA1 neurons at rest and during acute hypoxia. We recorded intracellularly from CA1 neurons in hippocampal slices, monitored V_m and measured input resistance (R_m) with periodic injections of negative current. We found that tetrodotoxin (TTX, 1 μM) hyperpolarized CA1 neurons at rest and significantly attenuated both the rate of depolarization (ΔV_m/dt) and the rate of decline of R_m (ΔR_m/dt) by about 60% during the early phase of hypoxia. The effect of TTX was dose-dependent. Amiloride (1 mM) decreased V_m and increased R_m in the resting condition but changed little the effect of hypoxia on neuronal function. Benzamil and 5-(N-ethyl-N-isopropyl)-2′,4′-amiloride (EIPA), two specific inhibitors of Na⁺ dependent exchangers, were similar to amiloride in their effect. We conclude that neuronal membrane properties are better maintained during anoxia by reducing the activity of TTX-sensitive channels and not by the action of Na⁺-dependent exchangers.     

Layer-specific mitochondrial diversity across hippocampal CA2 dendrites

CA2 is an understudied subregion of the hippocampus that is critical for social memory. Previous studies identified multiple components of the mitochondrial calcium uniporter (MCU) complex as selectively enriched in CA2. The MCU complex regulates calcium entry into mitochondria, which in turn regulates mitochondrial transport and localization to active synapses. We found that MCU is strikingly enriched in CA2 distal apical dendrites, precisely where CA2 neurons receive entorhinal cortical input carrying social information. Furthermore, MCU-enriched mitochondria in CA2 distal dendrites are larger compared to mitochondria in CA2 proximal apical dendrites and neighboring CA1 apical dendrites, which was confirmed in CA2 with genetically labeled mitochondria and electron microscopy. MCU overexpression in neighboring CA1 led to a preferential localization of MCU in the proximal dendrites of CA1 compared to the distal dendrites, an effect not seen in CA2. Our findings demonstrate that mitochondria are molecularly and structurally diverse across hippocampal cell types and circuits, and suggest that MCU can be differentially localized within dendrites, possibly to meet local energy demands.     

Modulation of 5HT1A Responsiveness in CA1 Pyramidal Neurons by in vivo Activation of Corticosteroid Receptors

In this study we describe modulatory effects exerted by in vivo activation of corticosteroid receptors on 5HT responsiveness of rat CA1 pyramidal neurons. In the first series of experiments, adrenalectomized (ADX) rats were injected with corticosterone one hour prior to decapitation (1–1000  μg/100  g body weight) after which 5HT_1A induced hyperpolarizations were determined in vitro by means of intracellular recordings. It appeared that 5HT responsiveness was dose-dependently affected by corticosterone injections: 5HT responses were relatively large when no corticosteroid receptors were activated (ADX); similar 5HT responses were observed or when both mineralocorticoid receptors (MR) and glucocorticoid receptors (GR) were occupied by injection of high doses of corticosterone (100–1000  μg/100  g body weight). However, compared to the latter group, 5HT hyperpolarizations were significantly suppressed in slices from rats that received moderate amounts of corticosterone (10–30  μg/100g). Next, we investigated whether physiological variations of plasma corticosterone levels as occurring in intact rats correlated with the transmitter responsiveness. It was found that high plasma levels of corticosterone due to either stress or exogenous application of high doses of corticosterone correlated with large 5HT-responses in vitro . Interestingly, the large 5HT responses recorded after stress were clearly suppressed by pretreatment with RU38486, a GR antagonist. Altogether, this study presents further evidence that 5HT transmission in hippocampal CA1 area is modulated by differential steroid receptor activation as may occur under physiological circumstances due to different plasma concentrations of corticosterone.     

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.