Chaotic dynamics as a mechanism of rapid transition of hippocampal local field activity between theta and non-theta states.

We propose a dynamical model of the local hippocampal circuit realizing the transition between the theta and non-theta states. We model the interaction between hippocampal local rhythm generators and the external periodic input from the medial septum and diagonal band of Broca (MS-DBB). With our model, bifurcation of the nonlinear dynamics serves as a mechanism that realizes two distinctive oscillations in the hippocampus, where the amplitude of the oscillatory input from the MS-DBB works as a bifurcation parameter. We model the network of the hippocampal interneurons with a network of simple class 1 neuron models connected mutually with gap junctions. The model neurons exhibit highly synchronous periodic oscillations under the existence of an external force from the MS-DBB, just as the real hippocampus shows theta oscillation under the rhythmic input from the MS-DBB. The model shows diffusion-induced chaotic dynamics under an aperiodic MS-DBB activity, just as the large amplitude irregular activity appears following the disappearance of the rhythmicity of the MS-DBB neurons in the real brain. The model is consistent with both previous experimental findings reporting the existence of local rhythm generators in the hippocampus and the executive role of the MS-DBB in synchronizing theta oscillation in vivo. Our model also replicates the traveling waves of theta oscillations in two-dimensionally coupled networks.

Layer- and cell type-selective co-transmission by a basal forebrain cholinergic projection to the olfactory bulb.

Cholinergic neurons in the basal forebrain project heavily to the main olfactory bulb, the first processing station in the olfactory pathway. The projections innervate multiple layers of the main olfactory bulb and strongly influence odor discrimination, detection, and learning. The precise underlying circuitry of this cholinergic input to the main olfactory bulb remains unclear, however. Here, we identify a specific basal forebrain cholinergic projection that innervates select neurons concentrated in the internal plexiform layer of the main olfactory bulb. Optogenetic activation of this projection elicits monosynaptic nicotinic and GABAergic currents in glomerular layer-projecting interneurons. Additionally, we show that the projection co-expresses markers for GABAergic neurotransmission. The data thus implicate neurotransmitter co-transmission in the basal forebrain regulation of this inhibitory olfactory microcircuit.Cholinergic neurons innervate multiple layers in the main olfactory bulb but the precise circuitry of this input is not known. Here the authors show that VGLUT3+ cholinergic neurons selectively innervate deep short axon cells in specific layers and elicit robust monosynaptic GABAergic and nicotinic postsynaptic currents.

Cognitive Deficits Associated with Nav1.1 Alterations: Involvement of Neuronal Firing Dynamics and Oscillations.

Brain oscillations play a critical role in information processing and may, therefore, be essential to uncovering the mechanisms of cognitive impairment in neurological disease. In Dravet syndrome (DS), a mutation in SCN1A, coding for the voltage-gated sodium channel Nav1.1, is associated with severe cognitive impairment and seizures. While seizure frequency and severity do not correlate with the extent of impairment, the slowing of brain rhythms may be involved. Here we investigate the role of Nav1.1 on brain rhythms and cognition using RNA interference. We demonstrate that knockdown of Nav1.1 impairs fast- and burst-firing properties of neurons in the medial septum in vivo. The proportion of neurons that fired phase-locked to hippocampal theta oscillations was reduced, and medial septal regulation of theta rhythm was disrupted. During a working memory task, this deficit was characterized by a decrease in theta frequency and was negatively correlated with performance. These findings suggest a fundamental role for Nav1.1 in facilitating fast-firing properties in neurons, highlight the importance of precise temporal control of theta frequency for working memory, and imply that Nav1.1 deficits may disrupt information processing in DS via a dysregulation of brain rhythms.

Hippocampal acetylcholine depletion has no effect on anxiety, spatial novelty preference, or differential reward for low rates of responding (DRL) performance in rats.

We investigated the role of the septo-hippocampal cholinergic projection in anxiety, spatial novelty preference, and differential reward for low rates of responding (DRL) performance. Cholinergic neurons of the rat medial septum (MS) and the vertical limb of the diagonal band of Broca (VDB) were lesioned using the selective immunotoxin, 192 IgG-saporin. Rats were then tested on several behavioral tests previously shown to be sensitive to either (a) hippocampal lesions or (b) nonselective MS/VDB lesions which target both cholinergic and γ-aminobutyric acid (GABA)-ergic projections, or both. Saporin lesions substantially reduced hippocampal cholinergic innervation, resulting in an absence of acetyl cholinesterase staining and markedly reduced choline acetyltransferase activity (mean reduction: 80 ± 5%; range: 50-97%). However, the saporin-lesioned rats did not differ from control rats in any of the behavioral tests. Thus we found no evidence from these lesion studies that the septo-hippocampal cholinergic projection plays an essential role in anxiety, spatial novelty preference, or DRL.

Interconnection and synchronization of neuronal populations in the mouse medial septum/diagonal band of Broca.

The medial septum/diagonal band of Broca (MS/DBB) is crucial for hippocampal theta rhythm generation (4-12 Hz). However, the mechanisms behind theta rhythmogenesis are still under debate. The MS/DBB consists, in its majority, of three neuronal populations that use acetylcholine, GABA, or glutamate as neurotransmitter. While the firing patterns of septal neurons enable the MS/DBB to generate rhythmic output critical for the generation of the hippocampal theta rhythm, the ability to synchronize these action potentials is dependent on the interconnectivity between the three major MS/DBB neuronal populations, yet little is known about intraseptal connections. Here we assessed the connectivity between pairs of MS/DBB neurons with paired patch-clamp recordings. We found that glutamatergic and GABAergic neurons provide intraseptal connections and produce sizable currents in MS/DBB postsynaptic cells. We also analyzed linear and nonlinear relationships between the action potentials fired by pairs of neurons belonging to various MS/DBB neuronal populations. Our results show that while the synchrony index for action potential firing was significantly higher in pairs of GABAergic neurons, coherence of action potential firing in the theta range was similarly low in all pairs analyzed. Recurrence analysis demonstrated that individual action potentials were more recurrent in cholinergic neurons than in other cell types. Implementing sparse connectivity in a computer model of the MS/DBB network reproduced our experimental data. We conclude that the interplay between the intrinsic membrane properties of different MS/DBB neuronal populations and the connectivity among these populations underlie the ability of the MS/DBB network to critically contribute to hippocampal theta rhythmogenesis.

Damage of GABAergic neurons in the medial septum impairs spatial working memory and extinction of active avoidance: effects on proactive interference.

The medial septum and diagonal band (MSDB) are important in spatial learning and memory. On the basis of the excitotoxic damage of GABAergic MSDB neurons, we have recently suggested a role for these neurons in controlling proactive interference. Our study sought to test this hypothesis in different behavioral procedures using a new GABAergic immunotoxin. GABA-transporter-saporin (GAT1-SAP) was administered into the MSDB of male Sprague-Dawley rats. Following surgery, rats were trained in a reference memory water maze procedure for 5 days, followed by a working memory (delayed match to position) water maze procedure. Other rats were trained in a lever-press avoidance procedure after intraseptal GAT1-SAP or sham surgery. Intraseptal GAT1-SAP extensively damaged GABAergic neurons while sparing most cholinergic MSDB neurons. Rats treated with GAT1-SAP were not impaired in acquiring a spatial reference memory, learning the location of the escape platform as rapidly as sham rats. In contrast, GAT1-SAP rats were slower than sham rats to learn the platform location in a delayed match to position procedure, in which the platform location was changed every day. Moreover, GAT1-SAP rats returned to previous platform locations more often than sham rats. In the active avoidance procedure, intraseptal GAT1-SAP impaired extinction but not acquisition of the avoidance response. Using a different neurotoxin and behavioral procedures than previous studies, the results of this study paint a similar picture that GABAergic MSDB neurons are important for controlling proactive interference.

Network effects of glutamate on neuronal activity in the medial septum/diagonal band complex in vitro.

Inter-neuronal interactions within the medial septum/diagonal band complex (MSDB) are of great interest as this region is believed to be the hippocampal theta rhythm pacemaker. However, the role of glutamatergic system in functioning of the septal cells is yet unclear. Here, we demonstrate for the first time the effects of glutamate in physiological concentration (1 microM) on the MSDB neuronal spontaneous and evoked activities in vitro. These effects (activation of 70% and inhibition of 30% of responsive neurons) differed in pacemaker and non-pacemaker cells. Pacemaker cells were always activated under glutamate, whereas non-pacemaker neurons could be either activated or inhibited. Indeed, in the burst pacemakers, glutamate increased the frequency of rhythmic activity. In a total MSDB neuron population, in 30% of neurons glutamate applications modified responses to the electrical stimulation by unifying the temporal parameters of neuron responses. Along with the increase in the theta-burst frequency, this indicates that the glutamatergic system is involved in the process ofintraseptal synchronization. Obtained data shed light on the role ofglutamatergic system in septal neuron interactions and broaden our understanding of theta oscillation mechanisms in the septo-hippocampal system.

Immunolocalization of the voltage-gated potassium channel Kv2.2 in GABAergic neurons in the basal forebrain of rats and mice.

The Kv2 voltage-gated potassium channels, Kv2.1 and Kv2.2, are important regulators of neuronal excitability in mammalian brain. It has been shown that Kv2.1 channels are expressed in virtually all neurons in the brain. However, the cellular localization of Kv2.2 has not been fully elucidated. In this article we report that Kv2.2 is highly expressed in a subset of neurons in the magnocellular preoptic nucleus (MCPO) and the horizontal limb of the diagonal band of Broca (HDB) of the basal forebrain complex, which are areas highly implicated in the regulation of cortical activity and the sleep/wake cycle. It has been shown that MCPO and HDB contain distinct populations of neurons that differ in their neurochemicals, cholinergic, glutamatergic, and gamma-aminobutyric acid (GABA)ergic neurons. Using specific immunolabeling and knockin mice in which green fluorescent protein (GFP) is expressed in GABAergic neurons, we found that Kv2.2 is abundantly expressed in a large subpopulation of the GABAergic neurons in the MCPO and HDB. These data offer Kv2.2 as a molecular target to study the role of the specific subpopulation of basal forebrain GABAergic neurons.

Distribution and role of Kv3.1b in neurons in the medial septum diagonal band complex.

The medial septum diagonal band complex (MS/DB) projects via cholinergic and GABAergic pathways to the hippocampus and plays a key role in the hippocampal theta rhythm. In the MS/DB we have previously described a population of fast spiking GABAergic neurons that contain parvalbumin and mediate theta frequency activity in vitro. The Kv3.1 potassium channel is a delayed rectifier channel that plays a major role in fast spiking neurons in the CNS, and has previously been localized in the MS/DB. To determine which cell types in the MS/DB express the Kv3.1b ion channel subunit, transgenic mice in which the expression of GABAergic and glutamate markers are associated with the expression of green fluorescent protein (GFP; GAD67-GFP and VGluT2-GFP mice, respectively) were used for immunofluorescence and axonal tract tracing. Electrophysiological studies were also carried out on rat MS/DB slices to examine the role of the Kv3.1 channel in theta frequency oscillations. The results for the MS/DB were as follows: (1) cholinergic cells did not express GFP in either GAD67-GFP or VGluT2-GFP mice, and there was GAD67 immunoreactivity in GFP-positive neurons in GAD67-GFP mice and in a small proportion (6%) of GFP-positive neurons in VGluT2-GFP mice. (2) Kv3.1b immunofluorescence was associated with the somata of GABAergic neurons, especially those that contained parvalbumin, and with a minority of glutamatergic neurons, but not with cholinergic neurons, and with GABAergic axonal terminal-like processes around certain GABAergic neurons. (3) Both Kv3.1b-positive and -negative GABAergic neurons were septo-hippocampal, and there was a minor projection to hippocampus from VGluT2-GFP neurons. (4) Kainate-induced theta oscillations in the MS/DB slice were potentiated rather than inhibited by the Kv3.1 blocker 4-aminopyridine, and this agent on its own produced theta frequency oscillations in MS/DB slices that were reduced by ionotropic glutamate and GABA receptor antagonists and abolished by low extracellular calcium. These studies confirm the presence of heterogeneous populations of septo-hippocampal neurons in the MS/DB, and suggest that presence of Kv3.1 in the GABAergic neurons does not contribute to theta activity through fast spiking properties, but possibly by the regulation of transmitter release from axonal terminals.

Excitatory effects of the puberty-initiating peptide kisspeptin and group I metabotropic glutamate receptor agonists differentiate two distinct subpopulations of gonadotropin-releasing hormone neurons.

Activation of the G-protein-coupled receptor GPR54 by kisspeptins during normal puberty promotes the central release of gonadotropin-releasing hormone (GnRH) that, in turn, leads to reproductive maturation. In humans and mice, a loss of function mutations of GPR54 prevents the onset of puberty and leads to hypogonadotropic hypogonadism and infertility. Using electrophysiological, morphological, molecular, and retrograde-labeling techniques in brain slices prepared from vGluT2-GFP and GnRH-GFP mice, we demonstrate the existence of two physiologically distinct subpopulations of GnRH neurons. The first subpopulation is comprised of septal GnRH neurons that colocalize vesicular glutamate transporter 2 and green fluorescent protein and is insensitive to metabotropic glutamate receptor agonists, but is exquisitely sensitive to kisspeptin which closes potassium channels to dramatically initiate a long-lasting activation in neurons from prepubertal and postpubertal mice of both sexes. A second subpopulation is insensitive to kisspeptin but is uniquely activated by group I metabotropic glutamate receptor agonists. These two physiologically distinct classes of GnRH cells may subserve different functions in the central control of reproduction and fertility.

The hippocamposeptal pathway generates rhythmic firing of GABAergic neurons in the medial septum and diagonal bands: an investigation using a complete septohippocampal preparation in vitro.

The medial septum diagonal band area (MS/DB) projects to the hippocampus through the fornix/fimbria pathway and is implicated in generating hippocampal theta oscillations. The hippocampus also projects back to the MS/DB, but very little is known functionally about this input. Here, we investigated the physiological role of hippocamposeptal feedback to the MS/DB in a complete in vitro septohippocampal preparation containing the intact interconnecting fornix/fimbria pathway. We demonstrated that carbachol-induced rhythmic theta-like hippocampal oscillations recorded extracellularly were synchronized with powerful rhythmic IPSPs in whole-cell recorded MS/DB neurons. Interestingly, we found that these IPSPs evoked rebound spiking in GABAergic MS/DB neurons. In contrast, putative cholinergic and glutamatergic MS/DB neurons responded only weakly with rebound spiking and, as a result, were mostly silent during theta-like oscillations. We next determined the mechanism underlying the rebound spiking that followed the IPSPs in MS/DB GABAergic neurons using phasic electrical stimulation of the fornix/fimbria pathway. We demonstrate that the increased rebound spiking was attributable to the activation of I(h) current, because it was significantly reduced by low concentrations of the I(h) antagonist ZD7288 [4-(N-ethyl-N-phenylamino)-1,2-dimethyl-6-(methylamino) pyridinium chloride]. Together, these results suggest that rhythmical activity in hippocampus is transferred to the MS/DB and can preferentially phase the spiking of GABAergic MS/DB neurons because of their significant expression of I(h) currents. Our data demonstrate that hippocamposeptal inhibition facilitates theta rhythmic discharges in MS/DB GABAergic neurons while favoring the inhibition of most ACh and glutamate neurons.

Postnatal lead exposure alters expression of forebrain p75 and TrkA nerve growth factor receptors.

Lead is a potent developmental neurotoxicant that affects many aspects of cognition and behavior. The hippocampus and striatum are among the areas particularly sensitive to the effects of lead and cholinergic neurons in both regions depend upon nerve growth factor (NGF) for their survival and maturation. The present study examined the extent to which postnatal lead exposure may affect the survival and expression of neuroptrophin receptors of septo-hippocampal cholinergic projection neurons in the medial septum/vertical limb of the diagonal band of Broca (MS/VDB) and cholinergic neurons of the striatum. Weanling rats were fed chow containing lead acetate for 30 days and effects on cholinergic cell number and the number of cells expressing neurotrophin receptors p75(NGFR) and trkA were assessed. A decrease in the number of cells expressing p75(NGFR) and an increase in the number of cells expressing trkA receptor was observed in the MS/VDB of lead-exposed rats, without a loss of cholinergic cell number or alteration in cell size. Lead-exposure resulted in a significant decrease in trkA-expressing cells in the striatum but no change in the number or size of cholinergic neurons. These results suggest that a brief postnatal lead exposure does not result in loss of MS/VDB or striatal cholinergic neurons but does modify the expression of neurotrophin receptors in these regions. The significance of these effects on the septo-hippocampal and striatal functioning remains to be studied.

Amyloid beta protein modulates glutamate-mediated neurotransmission in the rat basal forebrain: involvement of presynaptic neuronal nicotinic acetylcholine and metabotropic glutamate receptors.

Amyloid beta (Abeta) protein, a 39-43 amino acid peptide deposited in brains of individuals with Alzheimer's disease (AD), has been shown to interact directly with a number of receptor targets including neuronal nicotinic acetylcholine receptors (nAChRs) and glutamate receptors. In this study, we investigated the synaptic effects of Abeta(1-42) on glutamate-mediated neurotransmission in the diagonal band of Broca (DBB), a cholinergic basal forebrain nucleus. Glutamatergic miniature EPSCs (mEPSCs) were recorded using whole-cell patch-clamp recordings from identified cholinergic DBB neurons in rat forebrain slices. In 54% of DBB neurons, bath application of Abeta(1-42) (100 nM), but not Abeta(42-1) (inverse fragment), significantly increased the frequency of mEPSCs without affecting amplitude or kinetic parameters (rise or decay time). In 32% of DBB neurons, bath application of Abeta(1-42) significantly decreased only the frequency but not amplitude of mEPSCs. Application of dihydro-beta-erythroidine (DHbetaE) (an antagonist for the alpha4beta2 subtype of nAChRs) but not alpha-bungarotoxin (an antagonist for the alpha7 subtype of nAChRs) blocked Abeta(1-42)-mediated increases in mEPSC frequency. The Abeta(1-42)-mediated increase in glutamatergic transmission is thus presynaptic and mediated via non-alpha7 AChRs. In contrast, Abeta(1-42)-mediated decreases in mEPSC frequency could not be antagonized by either DHbetaE or alpha-bungarotoxin. However, the Abeta(1-42)-evoked depression in mEPSC frequency was antagonized by (RS)-alpha-methyl-4-carboxyphenyglycine, a nonselective group I/II metabotropic glutamate receptor antagonist. These observations provide further insight into the mechanisms whereby Abeta affects synaptic function in the brain and may be relevant in the context of synaptic failure observed in AD.

Cholinergic lesions produce task-selective effects on delayed matching to position and configural association learning related to response pattern and strategy.

192IgG-saporin (SAP) was used to selectively destroy cholinergic neurons in the rostral basal forebrain (e.g., medial septum (MS) and vertical limb of the diagonal band of Broca (VDB)) and/or the caudal basal forebrain (e.g., nucleus basalis magnocellularis (NBM)) of ovariectomized Sprague-Dawley rats. The effects of these lesions on two different cognitive tasks, a delayed matching to position (DMP) T-maze task, and a configural association (CA) operant conditioning task, were evaluated and compared. Injecting SAP into either the MS or NBM significantly impaired acquisition of the DMP task. Analysis showed that the effects were due largely to an affect on response patterns adopted by the rats during training, as opposed to an effect on working memory performance. Notably, the impairment in DMP acquisition did not correlate with the degree of cholinergic denervation of the hippocampus. Despite the deficit, most animals eventually learned the task and reached criterion; however by the end of training, controls and animals that received SAP into either the MS or NBM appeared more likely to use an allocentric place strategy to solve the task, whereas animals that received SAP into both the MS and NBM were more likely to use an egocentric response strategy. Cholinergic lesions also produced a small but significant affect on acquisition of the CA task, but only with respect to response time, and only in the SAP-NBM-treated animals. SAP-NBM lesions also produced small but significant impairments in both the number of responses and response time during the acquisition of simple associations, possibly reflecting an effect on alertness or attention. Notably, the effects on CA acquisition were small, and like the effects on DMP acquisition did not correlate with the degree of cholinergic denervation of the hippocampus. We conclude that selective basal forebrain cholinergic lesions produce learning deficits that are task specific, and that cholinergic denervation of either the frontal cortex or hippocampus can affect response patterns and strategy in ways that affect learning, without necessarily reflecting deficits in working memory performance.

Interaction of GABA and glutamate in the horizontal limb of diagonal band of Broca (hDB): role in cardiovascular responses.

The diagonal band of Broca (DBB) is a part of the limbic system that consists of two parts: the vertical limb (vDB) and the horizontal limb (hDB). It has been shown that microinjection of glutamate into the hDB of the anesthetized rat elicited depressor responses. We have previously shown that microinjection of AP5 (an NMDA receptor antagonist, 2.5 mM, 50 nl) and CNQX (an AMPA receptor antagonist, 1 mM, 50 nl) caused no significant changes in the blood pressure and heart rate. Microinjection of bicuculline (BMI: a GABA(A) receptor antagonist, 1 mM, 50 nl) resulted in the increase of both the blood pressure and heart rate. In this study, we investigated the possible interaction of the GABAergic and glutaminergic systems of the hDB by coinjection of the antagonists of both systems. Experiments were performed on the 24 urethane-anesthetized rats. Repeated measures ANOVA was used for data analysis. Our results showed that coinjection of 50 nl of 1 mM of BMI and 2.5 mM of AP5 significantly (ANOVA, P < 0. 01) decreased the pressor effects of BMI. Also, coinjection of 50 nl of BMI (1 mM) and CNQX (1 mM) significantly (ANOVA, P < 0.01) decreased the pressor effects of BMI. Coinjection of the previous amounts of BMI and both of the glutamate receptor antagonists also produced the same results. These results showed that the cardiovascular effects of blockade of GABAergic inhibition in the hDB are dependent on the activation of local NMDA and non-NMDA receptors of glutamate. A possible interpretation of the results is that, the GABAergic neurons inhibit the glutaminergic neurons.

Nerve growth factor rapidly increases muscarinic tone in mouse medial septum/diagonal band of Broca.

Nerve growth factor (NGF) has been implicated in maintaining and regulating normal functioning of the septohippocampal pathway. However, many aspects of its physiological actions and the underlying mechanisms await elucidation. In this study, we investigated the effect of acute NGF exposure on neurons in the mouse medial septum/diagonal band of Broca (MS/DB), focusing on the cholinergic neurons and the subpopulation of noncholinergic neurons that were identified to be putatively GABAergic. We report that MS/DB neurons in a thin slice preparation, when exposed to NGF via bath perfusion, rapidly and indiscriminately increased the rate of spontaneous firing in all MS/DB neurons. However, focal application of NGF to individual MS/DB neurons increased spontaneous firing in cholinergic, but not in the noncholinergic, subpopulation. The NGF-induced effect on cholinergic neurons was direct, requiring activation and signaling via TrkA receptors, which were immunohistochemically localized to the cholinergic neurons in the MS/DB. TrkA receptors were absent in putative GABAergic MS/DB neurons, and blockade of TrkA signaling in these and other noncholinergic neurons had no effect on their firing activity after exposure to NGF. Conversely, methyl scopolamine, blocked the increased firing activity of noncholinergic neurons during bath perfusion of NGF. We propose a cell type-specific mode of action for NGF in the MS/DB. The neurotrophin directly enhances cholinergic neuronal activity in the MS/DB through TrkA-mediated signaling, increasing acetylcholine release and, thus, muscarinic tone. This increase in muscarinic tone, in turn, results in heightened firing activity in noncholinergic MS/DB neurons.

Alpha 7 nicotinic receptor subunit is present on serotonin neurons projecting to hippocampus and septum.

The serotonergic transmitter system regulates hippocampal activity through its raphe projection to hippocampus and medial septum/diagonal band of Broca complex (MS/DBB), and most likely also indirectly through its interaction with the cholinergic neurotransmitter system. Nicotine, e.g., enhances hippocampal serotonin release probably through presynaptic nicotinic receptors. We investigated the possible presence of the alpha 7-nicotinic subunit on serotonergic neurons projecting to hippocampus and MS/DBB. By retrograde neuronal tracing, hippocampal serotonergic neurons were identified and with double fluorescence immunostaining and Alexa-488 bound alpha-bungarotoxin the presence of active alpha 7 receptor on their soma was determined. Most of the retrogradely labeled serotonin neurons contained the alpha 7 subunit. A low degree of colocalization between alpha-bungarotoxin and serotonin-positive neurons suggest that the alpha 7 subunit may be transported anterogradely to the serotonergic axonal terminals.

5-HT1A receptor mRNA and immunoreactivity in the rat medial septum/diagonal band of Broca-relationships to GABAergic and cholinergic neurons.

Activation of 5-HT1A receptors results in a variety of physiological responses, depending on their localization on neurons with different phenotypes in the brain. This study investigated the localization of 5-HT1A receptor mRNA and 5-HT1A receptor immunoreactivity in cell bodies of the rat septal complex using in situ hybridization and immunohistochemistry. In adjacent sections of the medial septum/diagonal band of Broca (MSDB), the distribution of cell bodies expressing 5-HT1A receptor mRNA was closely related to cells labeled with oligonucleotide probes to GAD (glutamic acid decarboxylase), VAChT (vesicular acetylcholine transporter) or parvalbumin mRNA. Using antiserum to GAD and antibodies to GABA, 5-HT1A receptor immunoreactivity was demonstrated in a majority of GABAergic cells in the MSDB. 5-HT1A receptor-immunoreactive GABAergic cells in the MSDB were also demonstrated to contain the calcium-binding protein parvalbumin, a marker for septohippocampal projecting GABAergic neurons. In the lateral septum, 5-HT1A receptor immunoreactivity was colocalized with the calcium-binding protein calbindin D-28k, a marker for septal GABAergic somatospiny neurons. 5-HT1A receptor immunoreactivity was also detected in a subpopulation of VAChT-containing cholinergic neurons of the MSDB. In MSDB neurons, colocalization of 5-HT1A and 5-HT2A receptor immunoreactivities was demonstrated. These observations suggest that serotonin via 5-HT1A receptors may represent an important modulator of hippocampal transmission important for cognitive and emotional functions through actions on both GABAergic and cholinergic neurons of the rat septal complex. In addition, 5-HT may exert its effects in the MSDB via cells expressing both 5-HT1A and 5-HT2A receptors.

Habituation to stress and dexamethasone suppression in rats with selective basal forebrain cholinergic lesions.

Previous studies suggest a role for basal forebrain cholinergic neurons in enhancing the inhibitory influence of the hippocampus and medial prefrontal cortex (mPFC) on glucocorticoid stress responses mediated by the hypothalamic-pituitary-adrenocortical (HPA) axis. An inhibitory action of the basal forebrain cholinergic (BFC) system may occur through facilitation of stress-related information processing and maintenance of glucocorticoid receptor (GR) expression and negative feedback signaling in these target regions. The current study investigated the possibility that BFC input to the hippocampus contributes to habituation of the glucocorticoid response following repeated exposure to a stressor. Cholinergic lesions were made by microinjections of the immunotoxin 192 IgG-saporin into the medial septum/vertical limb of the diagonal band, and 3 weeks later rats were subjected to six daily sessions of restraint stress. Blood samples taken before, during and after acute stress revealed a significant increase in peak activation and protracted elevation of corticosterone in cholinergic lesioned rats. After 5 days of repeated stress, however, both groups habituated to the stressor, as indicated by similarly low corticosterone profiles throughout both the response and recovery period. Against that habituated background, rats were administered a dexamethasone challenge on day 6, so that feedback status could be examined. Dexamethasone-induced suppression of endogenous corticosterone before, during, and after stress was significantly attenuated in lesioned rats. The profile of dysfunction in glucocorticoid regulation after selective cholinergic lesions in young animals may be relevant to the adrenocortical hyperactivity and negative feedback deficits seen in conditions such as normal aging and Alzheimer's dementia, in which integrity of the basal forebrain cholinergic system is compromised.

Detection of nitric oxide synthase (NOS) immunoreactive neurons in the human septal area: a matter of method?

There is a remarkable discrepancy between biochemical and cell morphological findings with regard to the presence of NADPH diaphorase/neuronal nitric oxide synthase (NOS) in the primate septal area. Whereas considerable concentrations of neuronal nitric oxide synthase and high enzyme activities have been measured in postmortem human septal nuclei, histochemical studies were either unable to detect any nitric oxide synthase immunoreactivity in primate septal neurons, or found only a very few nitrergic neurons in this region. This study aimed to investigate the possible presence of nitrergic neurons in human the septal region in greater detail. After having studied a total of 16 postmortem human brains we conclude that the immunohistochemical demonstration of nitric oxide synthase in human septal neurons is largely dependent on the mode of tissue handling: in brain specimens which were fixed en-bloc with paraffin and embedded in paraplast, nitric oxide synthase immunoreactivity is barely detectable, whereas a satisfying immunostaining is obtained on free-floating frozen sections after an immersion-fixation with 4% paraformaldehyde and 0.5% glutaraldehyde, followed by sucrose protection of the specimens. We show herein that there are indeed nitric oxide synthase-containing neurons in the human septum, thus supporting results from previous biochemical studies.