Cholinergic Septohippocampal Research Articles

Moderate lesion of the rat cholinergic septohippocampal pathway increases hippocampal nerve growth factor synthesis: evidence for long-term compensatory changes?

Moderate lesions of the septohippocampal pathway by intraventricular infusions of ethylcholine aziridinium (AF64A) induced a dose-dependent decrease of hippocampal choline acetyltransferase (ChAT) activity, which partially recovered between 1 and 5 weeks after treatment. The cholinergic deficit was associated with an increase in nerve growth factor (NGF) mRNA only within the hippocampal dentate gyrus and hilus by maximally 51% and 111% 3 and 7 weeks after AF64A treatment, respectively, whereas no changes in brain-derived neurotrophic factor- and neurotrophin-3 mRNA were observed. The content of NGF protein transiently increased in the ventral part of the hippocampus 3 weeks after AF64A infusion but returned to control levels at 5 weeks. At that time, however, NGF content as well as ChAT activity were significantly increased in the septum, suggesting an increased utilization of NGF by the remaining cholinergic neurons. Thus, the present data provide correlative evidence for a critical role of endogenous NGF in neuroregeneration and plasticity of the cholinergic basal forebrain in case of incipient damage.

Distinct Profile of Working Memory Errors Following Acute or Chronic Disruption of the Cholinergic Septohippocampal Pathway

The behavioral effects of two amnestic treatments (intraseptal chlordiazepoxide (CDP) and intraventricular AF64A) were examined in a delayed-nonmatch-to-sample radial-arm maze (DNMTS) paradigm. The types of errors induced by these treatments in this working memory task were assessed to determine how acute and chronic disruptions of the medial septum affect different phases of working memory (encoding, maintenance, retrieval). Rats were initially trained to perform the DNMTS task with a 1-h delay imposed between the training and the testing sessions. The first experiment demonstrated that intraseptal injection of 30 nmoles of CDP did not produce state-dependent learning. Rats were injected immediately following training with CDP or artificial cerobrospinal fluid (CSF; drug vehicle) and then prior to testing with CDP or CSF. Injection of CDP immediately following training (CDP–CSF) impaired performance of the task regardless of whether CDP was also administered before the postdelay test (CDP–CDP). Rats infused with CDP only before the postdelay test (CSF–CDP) exhibited a proactive deficit characterized by intact retention of the predelay information (i.e., arms entered prior to the delay) but impaired performance on the postdelay component (arms entered only after the delay). These data indicate: (i) that state dependency does not explain the working memory deficits induced by intraseptal CDP; (ii) that pretest CDP disrupts the storage and utilization of working memory for current arm selections. The second experiment examined the behavioral effects induced by a permanent disruption of the cholinergic septohippocampal pathway produced by icv injection of the cholinotoxin AF64A. Rats were initially trained on the DNMTS task and then bilaterally injected icv with either AF64A (2.5 nmoles/side) or CSF. AF64A-treated rats exhibited a significant impairment of performance compared to CSF-treated controls. In contrast to the impairment exhibited by CDP-treated rats in Experiment 1, the performance of AF64A-treated rats displayed a deficit in the maintenance/retrieval of information acquired during RAM trainingandan impairment in ability to store current spatial information in working memory to guide postdelay testing performance. These studies demonstrate that acute and chronic disruptions of the septohippocampal pathway produce distinct profiles of cognitive impairment that should help to reveal the behavioral and neurobiological characteristics of spatial memory.

Retrograde degenerative changes in the substantia nigra pars compacta following an excitotoxic lesion of the striatum

The retrograde changes induced by an excitotoxic lesion of the striatum (Str) on the neurons in substantia nigra pars compacta (SNc) projecting to the neuron-depleted region were investigated in adult rats. The retrograde tracer Fluoro-Gold (FG) was injected bilaterally into the Str. 2 weeks later, the excitotoxic amino acid ibotenic acid (IA) was injected unilaterally into the same structure. At four different time points after the lesion (1 week and 1,2 and 3 months, respectively), the size of the FG-labelled cells and number of tyrosine hydroxylase (TH)-positive cells in the SNc were evaluated on the lesioned and control sides. Parallel groups of animals received suspension grafts of fetal striatal tissue into the lesioned striata. At 1 week and 1 month after lesion, there were no changes in cell size, number of TH-positive cells or number of FG-labelled cells expressing TH in the SNc. At 2 and 3 months, however, there was a significant 30% shrinkage of the FG-labelled SNc cells but no evident decrease in TH-positive cell number, or in the expression of the TH protein, on the lesioned side as compared with the non-lesioned control side. Striatal transplants placed into the lesioned Str did not counteract this effect. This finding that an axon-sparing lesion of target cells results in cell shrinkage but no cell loss of the neurons that project to the lesioned area is in line with that has been shown to occur after similar lesions in the cholinergic septohippocampal and basalo-cortical systems. These slowly developing atrophic changes, without cell loss, induced in the nigral dopamine neurons by removal of the striatal target neurons should provide a useful model for the in vivo evaluation of the effect of putative target-derived neurotrophic factors in the nigrostriatal dopamine system.

Induction of c-FOS immunoreactivity in the hippocampus following potassium stimulation

In microdialysis procedures high potassium ion concentrations are generally used to induce neurotransmitter release. However, the widespread effects, if any, of such a treatment have not been described. In order to establish a possible link between c-mfos expression and stimulating conditions for neurotransmitter release in microdialysis procedures we administered KCl (100mM) into the hippocampus. Proto-oncogene c-FOS-like immunoreactivity is upregulated in granule cells of the dentate gyrus, pyramidal cells of the hippocampus, cingulate, piriform and frontoparietal cortices at 2 h, but not 24 h after K + administration. Neither implantation of the probes nor perfusion with artificial cerebrospinal fluid resulted in similar patterns of c-FOS immunoreactivity. In addition, we investigated whether the impairment of the cholinergic septohippocampal pathway would modify the K +-induced expression of the immediate early gene c-fos in the hippocampus. The expression of c-fos induced by KCl was not altered in the animals with fimbria-fornix lesion despite the marked decrease in acetylcholine release in the hippocampus. Glutamate concentrations measured in the same superfusates showed that a significantly greater glutamate release occurs in denervated hippocampi. Furthermore, abolishment of seizure-like activity (induced by KCl) in anaesthetized animals did not alter expression of c-FOS immunoreactivity in the K +-stimulated hippocampi. The results from these studies confirm that most of the releasable acetylcholine of the hippocampus is linked to the fimbrial input and may suggest that c-FOS upregulation in this model does not respond to any cholinergic input from the medial septum via the fimbria-fornix.

Oligodendrocyte- and myelin-associated inhibitors of neurite outgrowth: Their involvement in the lack of CNS regeneration

Until now central nervous system (CNS) neurites have been thought to have little capacity for regeneration following a lesion. When allowed to grow into peripheral nervous system (PNS) grafts, however, lesioned CNS axons are known to regenerate. Recently, an inhibitory substrate effect of CNS myelin and oligodendrocytes has been discovered which could be directly involved in the lack of regeneration. In culture, neurite growth cones were shown to specifically arrest their movement when contacting oligodendrocyte processes. The inhibitory components were characterized as two proteins of 35 and 250 kDa. A specific monoclonal antibody was generated (IN-1) that could neutralize these inhibitory effects. The role of the inhibitors in CNS regeneration was investigated in young rats receiving lesions of the corticospinal tract and implanted with a source of IN-1 mAB or control mAB. Results showed clear regeneration to over 10 mm in 2–5 weeks in IN-1 mAB-treated animals, while no fibers were detected further than 1 mm caudal to the lesion in controls. A similar, highly significant enhancement of regeneration was also found for the cholinergic septohippocampal pathway and for the optic nerve. These results show that lesioned CNS neurons can regenerate in CNS tissue when specific myelin components are neutralized, thus demonstrating that these inhibitory components play a crucial role in the lack of CNS regeneration.

Compensatory elevation of acetylcholine synthesis in vivo by cholinergic neurons surviving partial lesions of the septohippocampal pathway

The present study characterized the effects of partial destruction of the cholinergic septohippocampal pathway on transmitter functions of surviving cholinergic neurons in the hippocampus. Partial and full fimbrial transections were performed, and 3 weeks after lesioning, cholinergic functions were assessed in vivo and in vitro. Hippocampal ChAT activity and the capacity of hippocampal slices to synthesize [3H]ACh in vitro decreased by 35% and 45%, respectively, following partial fimbrial lesions and by 68% and 85%, respectively, following full fimbrial lesions. [3H]ACh release from hippocampal slices in vitro was decreased by 57% and 87%, respectively, following partial and full fimbrial lesions. Partial lesions decreased high-affinity choline uptake into hippocampal synaptosomes by 52%. In contrast to the significant reductions in cholinergic parameters measured in vitro after partial fimbrial lesions, such partial lesions did not significantly alter in vivo measures of hippocampal cholinergic function. Levels of endogenous ACh and choline measured in the hippocampus following partial lesions were similar to that of control values. Also, the hippocampal content of newly synthesized [2H4]ACh and the [2H4]ACh synthesis rate were not significantly different from control values. However, following full fimbrial lesions, in vivo measures of hippocampal cholinergic function were decreased to a degree similar to that observed in vitro. Hippocampal levels of endogenous ACh and [2H4]ACh and the synthesis rate for [2H4]ACh were decreased by 73%, 72%, and 83%, respectively. These results suggest that, following partial destruction of afferent cholinergic fibers that innervate the hippocampal formation, residual cholinergic neurons are able to upregulate their capacity to synthesize and store ACh in vivo, thus compensating for lesion-induced losses of cholinergic neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

Long-term administration of mouse nerve growth factor to adult rats with partial lesions of the cholinergic septohippocampal pathway

Nerve growth factor (NGF), a neurotrophic factor acting on cholinergic neurons of the basal forebrain, has been proposed as a treatment for Alzheimer's disease. Experimental support for its pharmacological use is derived from short-term studies showing that intraventricular administration of NGF during 2–4 weeks protects cholinergic cell bodies from lesion-induced degeneration, stimulates synthesis of choline acetyltransferase, and improves various behavioral impairments. To investigate the consequences of long-term NGF administration, we tested whether cholinergic cell bodies are protected from lesion-induced degeneration and whether cholinergic axons are stimulated to regrow into the denervated hippocampus following fimbrial transections. We found that intraventricular injections of NGF twice a week for 5 months to adult rats resulted in extended protection of cholinergic cell bodies from lesion-induced degeneration and did not produce obvious detrimental effects on the animals. NGF treatment mildly stimulated growth of cholinergic neurites within the 2-mm area directly adjacent to the fimbrial lesion but it failed to induce significant homotypic growth of cholinergic neurites into the deafferented hippocampus.

Differential hippocampal and cortical cholinergic activation during the acquisition, retention, reversal and extinction of a spatial discrimination in an 8-arm radial maze by mice

Possible differentiation of the intervention of cholinergic septohippocampal and magnocellular forebrain (NBM) projections to cortex during learning and memory processes has been investigated directly using mice. High-affinity choline uptake velocities in the hippocampus and cortex were analyzed, in parallel, at various periods during the acquisition, over 8 days, as were the subsequent retention, reversal and extinction of a spatial discrimination in an 8-arm radial maze. Initial acquisition induced an immediate (30 s) and long-lasting (approx. 3 h) increase in mean hippocampal (+ 33%) and cortical (+ 23%) cholinergic activities. The time course of this activation was structure-dependent and correlations of hippocampal-cortical cholinergic activities showed large and consistent alterations as a function of time after training. Cholinergic activation in both brain regions was observed immediately following each daily training session with amplitudes which did not vary significantly in spite of a progressive daily increment in performance. Following acquisition mice were tested for retention, reversal and extinction: 30 s following the retention session, cholinergic activation was observed in both cortex and hippocampus, with magnitudes similar to those observed at the end of acquisition. However, in the reversal and extinction groups, a treatment-dependent attenuation of cholinergic activation was observed which was accompanied by a significant loss of correlation of cholinergic activity between these two brain regions. The results are discussed in relation to the concepts of reference and working memory and also to novelty, stress, arousal and frustrative non-reward. The data constitute direct experimental evidence for a differential involvement of cholinergic septohippocampal and NBM-cortical projections in learning and memory processes.

Hippocampal formation neurons code the level of activation of the cholinergic septohippocampal pathway

In these experiments, differing levels of activation of the cholinergic septohippocampal pathway, as reflected by shifts in type 2 slow wave theta frequency recorded in the hippocampal formation, were studied in urethane-anesthetized rats. The purpose was to determine whether the discharge rates of cells in the hippocampal formation varied in any systematic way with the frequency shifts produced during spontaneous theta and by stimulation of the dorsomedial-posterior hypothalamic region. Linear regression analysis revealed that a class of neurons called theta-on cells precisely coded the level of activation of the septohippocampal pathway through their discharge rates. Administration of atropine sulfate abolished this coding. A second class of neurons called theta-off cells were shown to have reciprocal relationships with theta-on cells. Implications for the sensorimotor model of hippocampal formation theta subsystems were discussed.

Ontogeny of passive avoidance in the rat: Effect of an early septal lesion

The retrograde changes induced by an excitotoxic lesion of the striatum (Str) on the neurons in substantia nigra pars compacta (SNc) projecting to the neuron-depleted region were investigated in adult rats. The retrograde tracer Fluoro-Gold (FG) was injected bilaterally into the Str. 2 weeks later, the excitotoxic amino acid ibotenic acid (IA) was injected unilaterally into the same structure. At four different time points after the lesion (1 week and 1,2 and 3 months, respectively), the size of the FG-labelled cells and number of tyrosine hydroxylase (TH)-positive cells in the SNc were evaluated on the lesioned and control sides. Parallel groups of animals received suspension grafts of fetal striatal tissue into the lesioned striata. At 1 week and 1 month after lesion, there were no changes in cell size, number of TH-positive cells or number of FG-labelled cells expressing TH in the SNc. At 2 and 3 months, however, there was a significant 30% shrinkage of the FG-labelled SNc cells but no evident decrease in TH-positive cell number, or in the expression of the TH protein, on the lesioned side as compared with the non-lesioned control side. Striatal transplants placed into the lesioned Str did not counteract this effect. This finding that an axon-sparing lesion of target cells results in cell shrinkage but no cell loss of the neurons that project to the lesioned area is in line with that has been shown to occur after similar lesions in the cholinergic septohippocampal and basalo-cortical systems. These slowly developing atrophic changes, without cell loss, induced in the nigral dopamine neurons by removal of the striatal target neurons should provide a useful model for the in vivo evaluation of the effect of putative target-derived neurotrophic factors in the nigrostriatal dopamine system.

Some behavioural effects of non-reward in an 8-arm radial maze

Possible differentiation of the intervention of cholinergic septohippocampal and magnocellular forebrain (NBM) projections to cortex during learning and memory processes has been investigated directly using mice. High-affinity choline uptake velocities in the hippocampus and cortex were analyzed, in parallel, at various periods during the acquisition, over 8 days, as were the subsequent retention, reversal and extinction of a spatial discrimination in an 8-arm radial maze. Initial acquisition induced an immediate (30 s) and long-lasting (approx. 3 h) increase in mean hippocampal (+ 33%) and cortical (+ 23%) cholinergic activities. The time course of this activation was structure-dependent and correlations of hippocampal-cortical cholinergic activities showed large and consistent alterations as a function of time after training. Cholinergic activation in both brain regions was observed immediately following each daily training session with amplitudes which did not vary significantly in spite of a progressive daily increment in performance. Following acquisition mice were tested for retention, reversal and extinction: 30 s following the retention session, cholinergic activation was observed in both cortex and hippocampus, with magnitudes similar to those observed at the end of acquisition. However, in the reversal and extinction groups, a treatment-dependent attenuation of cholinergic activation was observed which was accompanied by a significant loss of correlation of cholinergic activity between these two brain regions. The results are discussed in relation to the concepts of reference and working memory and also to novelty, stress, arousal and frustrative non-reward. The data constitute direct experimental evidence for a differential involvement of cholinergic septohippocampal and NBM-cortical projections in learning and memory processes.

Laminar distribution of hippocampal rhythmic slow activity (RSA) in the behaving rat: Current-source density analysis, effects of urethane and atropine

This study investigated the laminar distribution of rhythmic slow wave activity (RSA) in the dorsal hippocampus of the rat during running. Depth analyses of field EEG were performed by stepping the recording electrode in 82.5 μm increments and sampling RSA at each depth. One-dimensional current-source density (CSD) was calculated from the RSA profiles to enhance spatial resolution of current sources and sinks. Laminar analysis of power, coherence, and phase of RSA with respect to a stationary electrode in the stratum oriens of CA1 was performed with spectral methods. RSA waves in the CA1-dentate axis had power maxima at about the hippocampal fissures, hilus, outer molecular layer of the endal leaf of dentate gyrus and stratum oriens of CA1, in that order. A gradual shift of phase occurred in stratum radiatum of CA1. Large phase-shifts were found in both the endal and ectal leaves of the fascia dentata. A null zone and associated sudden phase-reversal of RSA were observed in stratum lucidum of CA3. Multiunit activity showed phase-locked modulation with RSA in the granule cell layer of the dendate gyrus and pyramidal cell layer of CA1, CA3, and subiculum. CSD analysis in the CA1-dentate axis revealed multiple source-link pairs. The sinks and sources showed cyclic changes with RSA, and were attributed to the rhythmic, but time-shifted, activity of hippocampal afferents from the septum and entorhinal cortex. The gradual phase-shift in CA1, and the configurational changes of RSA waves with depth, are explained by the summation of extracellular currents produced by time-delayed sink-source pairs (RSA dipoles). When the cholinergic septohippocampal path was blocked by atropine a null zone in the middle of stratum radiatum of CA1 occurred and the phase-shift of RSA became steeper. Under urethane anesthesis a null zone was present in the inner stratum radiatum associated with a sudden phase-reversal of RSA. Urethane reduced the power of RSA in the hilus and decreased the firing rate of the granule cells. It is suggested that field RSA is produced by several rhythmical dipoles along the somadendritic surface of pyramidal cells and granule cells and the spatiotemporal relations of the individuals dipoles determine the actually observed extracellular RSA.

Modulation of inhibition in the hippocampus in vivo.

The nature and mechanisms of septohippocampal transmission have been elucidated by taking advantage of an in situ preparation in experiments with Sprague-Dawley rats under urethane. Both extracellular field potentials and intracellular recordings were made in CA1-3 regions of the hippocampus; and the hippocampal commissure and medial septum stimulated to evoke synaptic activity. Using muscarinic and nicotinic agonists and antagonists it was shown that both acetylcholine and medial septal activity can increase the excitability of pyramidal cells, mainly through muscarinic receptors. The effect of septal stimulation was enhanced by local application of physostigmine and reduced by intraventricular injections of hemicholinium. It was also shown that acetylcholine, when applied in the stratum pyramidale, can reduce the voltage and conductance changes observed during evoked inhibitory postsynaptic potentials (IPSP) without affecting the action of gamma-aminobutyric acid on membrane conductance and voltage. It is therefore proposed that acetylcholine can reduce evoked IPSPs through presynaptic inhibition. Evidence is also presented that medial septal stimulation can reduce the efficacy of evoked IPSPs. These observations provide further support for the existence of a cholinergic septohippocampal pathway.

Cells of origin of the ventral cholinergic septohippocampal pathway undergoing compensatory collateral sprouting following fimbria-fornix transection

The axons of the septohippocampal pathway reach the hippocampal formation via the fimbria, dorsal fornix and supracallosal striae. Complete lesions of these pathways denervate most of the hippocampal formation but a small, residual innervation occurs via a different ventral route. The cells of origin for this additional route for cholinergic innervation of the hippocampal formation have been identified in the present study with a combined technique using a fluorescent retrograde tracer and acetylcholinesterase histochemistry. These cells were localized to the diagonal band of Broca. In a subsequent experiment it was demonstrated, by destroying these cells unilaterally, that they were responsible to a large extent for the cholinergic compensatory collateral sprouting in the hippocampus that has previously been shown to develop following complete fimbria-fornix transection.

Sustained elevation of hippocampal cyclic 3'-5' adenosine monophosphate levels after medial septal lesions.

The cyclic 3'-5' adenosine monophosphate (cyclic AMP) content of the rat hippocampal formation doubles during the week following a medial septal lesion and remains elevated for at least 1 month, the longest time period studied. This elevation in cyclic AMP does not result from sympathetic ingrowth, as neither superior cervical ganglion stimulation nor ganglionectomy influences hippocampal cyclic AMP content after lesions. Interruption of the cholinergic septohippocampal pathway in the fornix did not elevate hippocampal cyclic AMP content. Further, treatment of septal-lesioned animals with oxotremorine or of normal animals with atropine did not influence hippocampal cyclic AMP content. Finally, neither locus ceruleus lesions nor treatment with propranolol affected hippocampal cyclic AMP content. We believe this to be the first report of a sustained elevation in hippocampal cyclic AMP content. Like other long-term events, it is likely to have profound effects on hippocampal function and represents a remarkable brain adaptation to remote injury.

An investigation of whether septal γ-aminobutyrate-containing interneurons are involved in the reduction in the turnover rate of acetylcholine elicited by substance P and β-endorphin in the hippocampus

Afferents to the septum that contain substance P or β-endorphin probably have an inhibitory action upon the acetylcholine metabolism of the cholinergic septohippocampal pathway because intraseptal administration of these peptides decreases the turnover rate of acetylcholine in the hippocampus. Bicuculline, an antagonist of γ-aminobutyric acid, blocks the inhibitory action of β-endorphin on acetylcholine turnover suggesting that it is mediated via the septal γ-aminobutyric acid-containing interneurons. Inhibition by substance P of the turnover of acetycholine in the hippocampus is independent of this type of interneuron. Dopamine, like β-endorphin, acts on γ-aminobutyrie acid-containing interneurons of the septum. However, lesions of dopaminergic cell bodies located in area A 10 have established that the dopaminergie and β-endorphin-containing inputs to the septal γ-aminobutyric acidcontaining interneurons are independent of each other.