Increase In Acetylcholine Efflux Research Articles

Cholinergic neurotransmission in the basolateral amygdala during cued fear extinction

Cholinergic neuromodulation plays an important role in numerous cognitive functions including regulating arousal and attention, as well as associative learning and extinction processes. Further, studies demonstrate that cholinergic inputs from the basal forebrain cholinergic system influence physiological responses in the basolateral amygdala (BLA) as well as fear extinction processes. Since rodent models display individual differences in conditioned fear and extinction responses, this study investigated if cholinergic transmission in the BLA during fear extinction could contribute to differences between extinction resistant and extinction competent phenotypes in outbred Long-Evans male rats. Experiment 1 used in vivo microdialysis to test the hypothesis that acetylcholine (ACH) efflux in the BLA would increase with presentation of an auditory conditioned stimulus (CS+) during extinction learning. Acetylcholine efflux was compared in rats exposed to the CS+, a CS- (the tone never paired with a footshock), or to a context shift alone (without CS+ tone presentation). Consistent with acetylcholine's role in attention and arousal, ACH efflux in the BLA was increased in all three groups (CS+, CS-, Shift Alone) by the initial context shift into the extinction learning chamber, but returned more rapidly to baseline levels in the Shift Alone group (no CS+). In contrast, in the group exposed to the CS+, ACH efflux in the BLA remained elevated during continued presentation of conditioned cues and returned to baseline more slowly, leading to an overall increase in ACH efflux compared with the Shift Alone group. Based on the very dense staining in the BLA for acetylcholinesterase (ACHE), Experiment 2 examined if individual differences in fear extinction were associated with differences in cholinesterase enzyme activity (CHE) in the BLA and/or plasma with a separate cohort of animals. Cholinesterase activity (post-testing) in both the BLA and plasma was higher in extinction competent rats versus rats resistant to extinction learning. There was also a significant negative correlation between BLA CHE activity and freezing during extinction learning. Taken together, our results support a role for ACH efflux in the BLA during cued fear extinction that may be modulated by individual differences in ACHE activity, and are associated with behavioral responses during fear extinction. These findings implicate individual differences in cholinergic regulation in the susceptibility to disorders with dysregulation of extinction learning, such post-traumatic stress disorder (PTSD) in humans.

Acetylcholine efflux from retrosplenial areas and hippocampal sectors during maze exploration

Both the retrosplenial cortex (RSC) and the hippocampus are important for spatial learning across species. Although hippocampal acetylcholine (ACh) release has been associated with learning on a number of spatial tasks, relatively little is understood about the functional role of ACh release in the RSC. In the present study, spatial exploration was assessed in rats using a plus maze spontaneous alternation task. ACh efflux was assessed simultaneously in the hippocampus and two sub-regions of the RSC (areas 29ab and 30) before, during and after maze exploration. Results demonstrated that there was a significant rise in ACh efflux in RSC area 29ab and the hippocampus during maze traversal. The rise in ACh efflux across these two regions was correlated. There were no significant behaviorally driven changes in ACh efflux in RSC area 30. While both the hippocampal sectors and area 29ab displayed increases in ACh efflux during maze exploration, the percent ACh rise in area 29ab was higher than that observed in the hippocampus and persisted into the post-baseline period. Joint efflux analyses demonstrated a key functional role for ACh release in area 29ab during spatial processing.

Effect of muscarinic receptor agonists xanomeline and sabcomeline on acetylcholine and dopamine efflux in the rat brain; comparison with effects of 4-[3-(4-butylpiperidin-1-yl)-propyl]-7-fluoro-4 H-benzo[1,4]oxazin-3-one (AC260584) and N-desmethylclozapine

We have demonstrated that the main metabolite of clozapine, N-desmethylclozapine, has a significant role in the ability of clozapine to improve some aspects of cognition in schizophrenia. Furthermore, there is also evidence to suggest that it is the muscarinic M 1 receptor agonist effect of N-desmethylclozapine that underlies its cognitive effects. In the present study we examined the efficacy of two muscarinic receptor agonists xanomeline and sabcomeline to increase the efflux of acetylcholine and dopamine in rat medial prefrontal cortex and nucleus accumbens. Microdialysis in awake, freely moving rats was used to demonstrate that xanomeline at 10, but not 1 or 3 mg/kg (s.c.), significantly increased acetylcholine efflux in both the medial prefrontal cortex and nucleus accumbens. Sabcomeline, at 1 but not 0.1 or 0.5 mg/kg (s.c.), significantly increased acetylcholine efflux in the medial prefrontal cortex but not the nucleus accumbens. Both xanomeline and sabcomeline dose-dependently increased dopamine efflux in the medial prefrontal cortex but only high dose of xanomeline (10 mg/kg, s.c.) and sabcomeline (1 mg/kg, s.c.) increased that in the nucleus accumbens. The acetylcholine and dopamine efflux induced by xamomeline (10 mg/kg, s.c.) and sabcomeline (1 mg/kg, s.c.) were significantly blocked by the preferential muscarinic M 1 receptor antagonist telenzepine (3 mg/kg, s.c.), but significantly potentiated by the atypical antipsychotic drug risperidone (0.1 mg/kg, s.c.), which does not have much affinity for muscarinic receptor(s). According to the analysis of net-AUC (area under the curve) values of acetylcholine and dopamine levels, the rank order of ability of these drugs to increase acetylcholine or dopamine levels is sabcomeline > xanomeline ≈ AC260584 > N-desmethylclozapine. The present study suggests that the binding potency of muscarinic M 1 receptors is greatly related to their ability to increase cortical acetylcholine and dopamine efflux, and that this may have some relevance for treatment of the cognitive deficit of schizophrenia.

Asenapine Increases Dopamine, Norepinephrine, and Acetylcholine Efflux in the Rat Medial Prefrontal Cortex and Hippocampus

Atypical antipsychotic drugs, which are more potent direct acting antagonists of brain serotonin (5-HT)(2A) than dopamine (DA) D(2) receptors, preferentially enhance DA and acetylcholine (ACh) efflux in the rat medial prefrontal cortex (mPFC) and hippocampus (HIP), compared with the nucleus accumbens (NAc). These effects may contribute to their ability, albeit limited, to improve cognitive function and negative symptoms in patients with schizophrenia. Asenapine (ASE), a new multireceptor antagonist currently in development for the treatment of schizophrenia and bipolar disorder, has complex serotonergic properties based upon relatively high affinity for multiple serotonin (5-HT) receptors, particularly 5-HT(2A) and 5-HT(2C) receptors. In the current study, the effects of ASE on DA, norepinephrine (NE), 5-HT, ACh, glutamate, and gamma-aminobutyric acid (GABA) efflux in rat mPFC, HIP, and NAc were investigated with microdialysis in awake, freely moving rats. ASE at 0.05, 0.1, and 0.5 mg/kg (s.c.), but not 0.01 mg/kg, significantly increased DA efflux in the mPFC and HIP. Only the 0.5 mg/kg dose enhanced DA efflux in the NAc. ASE, at 0.1 and 0.5 mg/kg, significantly increased ACh efflux in the mPFC, but only the 0.5 mg/kg dose of ASE increased HIP ACh efflux. ASE did not increase ACh efflux in the NAc at any of the doses tested. The effect of ASE (0.1 mg/kg) on DA and ACh efflux was blocked by pretreatment with WAY100635, a 5-HT(1A) antagonist/D(4) agonist, suggesting involvement of indirect 5-HT(1A) agonism in both the actions. ASE, at 0.1 mg/kg, increased NE, but not 5-HT, efflux in the mPFC and HIP. ASE, at 0.1 mg/kg (s.c.), had no effect on glutamate and GABA efflux in either the mPFC or NAc. These findings indicate that ASE is similar to clozapine and other atypical antipsychotic drugs in preferentially increasing the efflux of DA, NE, and ACh in the mPFC and HIP compared with the NAC, and suggests that, like these agents, it may also improve cognitive function and negative symptoms in patients with schizophrenia.

Selective septohippocampal – but not forebrain amygdalar – cholinergic dysfunction in diencephalic amnesia

A rodent model of diencephalic amnesia, pyrithiamine-induced thiamine deficiency (PTD), was used to investigate diencephalic–limbic interactions. In-vivo acetylcholine (ACh) efflux, a marker of memory-related activation, was measured in the hippocampus and the amygdala of PTD-treated and pair-fed (PF) control rats while they were tested on a spontaneous alternation task. During behavioral testing, all animals displayed increases in ACh efflux in both the hippocampus and amygdala. However, during spontaneous alternation testing ACh efflux in the hippocampus and the alternation scores were higher in PF rats relative to PTD-treated rats. In contrast, ACh efflux in the amygdala was not suppressed in PTD treated rats, relative to PF rats, prior to or during behavioral testing. In addition, unbiased stereological estimates of the number of choline acetyltransferase (ChAT) immunopositive neurons in the medial septal/diagonal band (MS/DB) and nucleus basalis of Meynert (NBM) also reveal a selective cholinergic dysfunction: In PTD-treated rats a significant loss of ChAT-immunopositive cells was found only in the MS/DB, but not in the NBM. Significantly, these results demonstrate that thiamine deficiency causes selective cholinergic dysfunction in the septo-hippocampal pathway.

Blunted hippocampal, but not striatal, acetylcholine efflux parallels learning impairment in diencephalic-lesioned rats

A rodent model of diencephalic amnesia, pyrithiamine-induced thiamine deficiency (PTD), was used to investigate the dynamic role of hippocampal and striatal acetylcholine (ACh) efflux across acquisition of a nonmatching-to-position (NMTP) T-maze task. Changes in ACh efflux were measured in rats at different time points in the acquisition curve of the task (early = day 1, middle = day 5, and late = day 10). Overall, the control group had higher accuracy scores than the PTD group in the latter sessions of NMTP training. During the three microdialysis sampling points, all animals displayed significant increases in ACh efflux in both hippocampus and striatum, while performing the task. However, on day 10, the PTD group showed a significant behavioral impairment that paralleled their blunted hippocampal—but not striatal—ACh efflux during maze training. The results support selective diencephalic–hippocampal dysfunction in the PTD model. This diencephalic–hippocampal interaction appears to be critical for successful episodic and spatial learning/memory.

Excitatory and Inhibitory Actions of Isoflurane on the Cholinergic Ascending Arousal System of the Rat

The cholinergic arousal systems are known to critically regulate the state of consciousness. The aim of this study was to determine the effect of isoflurane on the inhibitory or excitatory neurotransmitters efflux in important nuclei within the cholinergic arousal system using in vivo intracerebral microdialysis. The efflux of glutamate, gamma-aminobutyric acid (GABA), or acetylcholine in the posterior hypothalamus (PH), the basal forebrain (BF), and the somatosensory cortex (S1BF) of rats was detected using intracerebral microdialysis under an awake condition and at 0.5-2.0 minimum alveolar concentration (MAC) isoflurane anesthesia. The intrabasalis perfusion of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and N-methyl-D-aspartate on the cortical acetylcholine effluxes was also examined under both conditions. Isoflurane had no influence on the glutamate and GABA efflux in the PH, whereas in the BF, it dose-dependently increased glutamate efflux and decreased GABA efflux. A transient increase in glutamate efflux at 1.0 MAC and a decrease in GABA at 0.5-1.5 MAC were observed in the S1BF. Isoflurane dose-dependently decreased acetylcholine efflux in the S1BF. Perfusion of the BF with AMPA increased acetylcholine efflux in the S1BF with electroencephalographic activation during 0.75 MAC isoflurane anesthesia, suggesting an inhibitory action of isoflurane on AMPA receptors in the BF. However, N-methyl-D-aspartate had no effect on these parameters. Isoflurane induces both excitatory and inhibitory actions in the cholinergic arousal system. The predominant inhibitory action of isoflurane over its excitatory action at the BF would result in the decrease in the acetylcholine efflux in the S1BF.

Tolerance to the procholinergic action of the D1 receptor full agonist dihydrexidine

It has been well established that dopamine D1 receptor agonists increase acetylcholine (ACh) release in the brain, an effect that has been suggested to contribute to their procognitive properties. We sought to examine whether the ACh-releasing action of the D1 receptor full agonist dihydrexidine HCl (DHX) is altered after repeated administration. DHX dose-dependently (3, 9, 18, and 36 mg/kg, i.p.) increased ACh efflux in the hippocampus of freely moving rats through D1 receptor stimulation, as the D1 receptor antagonist SCH 23390 (0.3 mg/kg, i.p.) abolished the effect of DHX (18 mg/kg, i.p.). Daily injections of DHX (18 mg/kg, i.p.) over 14 days did not significantly affect basal hippocampal ACh concentrations, but they significantly curtailed the stimulatory action of a challenge injection of DHX (18 mg/kg, i.p.) on ACh efflux. Tolerance to the procholinergic action of DHX develops with repeated administration.

Augmented Prefrontal Acetylcholine Release during Challenged Attentional Performance

Previous research has demonstrated that attentional performance depends on the integrity of the cortical cholinergic input system and that such performance is associated with increases in cortical acetylcholine (ACh) release. The present experiment tested the hypothesis that the attentional impairments produced by bilateral basal forebrain infusions of the NMDA receptor antagonist DL-2-amino-5-phosphonovaleric acid (APV) are associated with attenuation of performance-associated increases in ACh release. Rats were trained in a sustained attention task and equipped with three guide cannula for the bilateral infusion of the NMDA receptor antagonist APV (0, 3, 20 nmol) and for the insertion of a dialysis probe into the medial prefrontal cortex (mPFC). APV or vehicle was infused remotely following completion of the first of five blocks of trials. During the first block, attentional performance was associated with a 140% increase in ACh efflux. Infusions of APV decreased the animals' ability to detect signals and augmented the increases in ACh efflux observed prior to infusions. These data indicate a dissociation between levels of attentional performance and increases in mPFC ACh release. Augmentation of performance-associated increases in mPFC cholinergic transmission is hypothesized to mediate the increased demands on attentional 'effort' that are required to maintain performance under challenging conditions.

Prefrontal cortical modulation of acetylcholine release in posterior parietal cortex

Attentional processing is a crucial early stage in cognition and is subject to “top-down” regulation by prefrontal cortex (PFC). Top-down regulation involves modification of input processing in cortical and subcortical areas, including the posterior parietal cortex (PPC). Cortical cholinergic inputs, originating from the basal forebrain cholinergic system, have been demonstrated to mediate important aspects of attentional processing. The present study investigated the ability of cholinergic and glutamatergic transmission within PFC to regulate acetylcholine (ACh) release in PPC. The first set of experiments demonstrated increases in ACh efflux in PPC following AMPA administration into the PFC. These increases were antagonized by co-administration of the AMPA receptor antagonist DNQX into the PFC. The second set of experiments demonstrated that administration of carbachol, but not nicotine, into the PFC also increased ACh efflux in PPC. The effects of carbachol were attenuated by co-administration (into PFC) of a muscarinic antagonist (atropine) and partially attenuated by the nicotine antagonist mecamylamine and DNQX. Perfusion of carbachol, nicotine, or AMPA into the PPC did not affect PFC ACh efflux, suggesting that these cortical interactions are not bi-directional. These studies demonstrate the capacity of the PFC to regulate ACh release in the PPC via glutamatergic and cholinergic prefrontal mechanisms. Prefrontal regulation of ACh release elsewhere in the cortex is hypothesized to contribute to the cognitive optimization of input processing.

Thalamic regulation of striatal acetylcholine efflux is both direct and indirect and qualitatively altered in the dopamine-depleted striatum

Striatal cholinergic interneurons play a pivotal role in the integrative sensorimotor functions of the basal ganglia. The major excitatory input to these interneurons arises from glutamatergic neurons of the parafascicular nucleus of the thalamus (Pf). Thalamic regulation of cholinergic interneurons, however, may also include an indirect inhibitory component mediated by the axon collaterals of GABAergic medium spiny neurons that are also innervated by Pf. The present study examined thalamic regulation of striatal cholinergic interneurons by employing dual probe in vivo microdialysis in freely moving animals to determine the effect of pharmacological manipulation of Pf on acetylcholine (ACh) efflux in intact and dopamine-lesioned striata. In intact animals, reverse dialysis application of the GABA A antagonist bicuculline (50 μM) into Pf, likely disinhibiting Pf neurons, significantly decreased striatal ACh efflux. When striatal GABA A receptors were blocked by simultaneous reverse dialysis application of bicuculline (10 μM), however, the same manipulation significantly increased ACh efflux. Qualitatively similar results were obtained in experiments employing a higher concentration of bicuculline (200 μM). Application of the GABA agonist muscimol (500 μM) into Pf, likely inhibiting Pf neurons, decreased ACh efflux only when the experiment was conducted under blockade of striatal GABA A receptors. These data are consistent with the existence of an indirect, inhibitory, GABA A receptor-mediated component of ACh regulation that is most clearly manifested when Pf is disinhibited and with the existence of a direct excitatory component of ACh regulation, evident when Pf is inhibited. Manipulation of Pf using very high concentrations of drug (500 μM bicuculline, 2 mM muscimol), however, yielded data consistent only with direct excitatory thalamic regulation. In contrast to results obtained in intact animals, in animals with prior (3 weeks) unilateral lesion of the dopaminergic nigrostriatal pathway, bicuculline application (50 μM) in Pf significantly increased striatal ACh efflux, irrespective of simultaneous blockade of striatal GABA A receptors. The results of experiments in which muscimol (500 μM) was applied in Pf were similar to those obtained in intact animals, however. Baseline ACh efflux was not significantly elevated in dopamine-lesioned animals. These results indicate a qualitative alteration in the effectiveness of an inhibitory component of the thalamic regulation of ACh efflux in the dopamine depleted striatum, evident during increased thalamostriatal input. Such altered regulation of striatal ACh output is likely to have profound consequences for integrative function in the parkinsonian basal ganglia.

The CB1 receptor antagonist SR141716A selectively increases monoaminergic neurotransmission in the medial prefrontal cortex: implications for therapeutic actions.

1. In order to explore potential therapeutic implications of cannabinoid antagonists, the effects of the prototypical cannabinoid antagonist SR141716A on monoamine efflux from the medial prefrontal cortex and the nucleus accumbens of the rat were investigated by in vivo microdialysis. 2. SR141716A moderately increased serotonin efflux and concentrations of its metabolite 5-HIAA, both in the medial prefrontal cortex and the nucleus accumbens, and increased norepinephrine, dopamine and their metabolites in the medial prefrontal cortex. In contrast, it had no effect on norepinephrine, dopamine and their metabolites in the nucleus accumbens. 3. At the same doses, SR141716A increased acetylcholine efflux in the medial prefrontal cortex, in agreement with previous studies; contrary to the effects in cortex, SR141716A had no effect on acetylcholine efflux in the nucleus accumbens. 4. The efficacy of SR141716A in the psychostimulant-induced hyperlocomotion and the forced swimming paradigms was also explored in mice. SR141716A attenuated phenylcyclidine- and d-amphetamine-induced hyperlocomotion, without affecting locomotor activity when administered alone, and decreased immobility in the forced swimming test. 5. These results suggest that the cortical selectivity in the release of catecholamines, dopamine in particular, induced by the cannabinoid antagonist SR141716A, its procholinergic properties, together with its mild stimulatory effects on serotonin and norepinephrine efflux make similar compounds unique candidates for the treatment of psychosis, affective and cognitive disorders.

The effects of manipulations of attentional demand on cortical acetylcholine release

In vivo microdialysis was used to measure acetylcholine (ACh) efflux in the frontoparietal cortex while rats performed in one of two operant tasks. One task was designed and validated to generate measures of sustained attention, while the other task was designed to minimize explicit demands on sustained attentional resources (low-demand task). Transferring animals from the baseline environment into the operant chambers robustly increased cortical ACh efflux regardless of subsequent task demands. Performance in the sustained attention task further increased frontoparietal ACh efflux, and these increases were not observed when animals were simply exposed to the operant chamber without task performance. Manipulations of the task parameters within a session, to either increase or decrease explicit demands on sustained attention, were not associated with fluctuations in ACh efflux. Unexpectedly, performance in the low-demand task was also associated with significant increases in ACh efflux that were similar to those observed during the sustained attention task. However, widespread depletions of cortical cholinergic inputs produced by intra-basalis infusions of 192 IgG-saporin failed to impair performance in the low-demand task, suggesting that cholinergic transmission is not necessary for performance in this task. The present results indicate that although a wider range of instrumental processes than previously hypothesized are associated with increases in cortical ACh release, the dependence of performance on the integrity of cortical cholinergic inputs may be limited to tasks with explicit attentional demands.

Intracranial self-stimulation induces Fos expression in GABAergic neurons in the rat mesopontine tegmentum

The cholinergic neurons which originate in the mesopontine tegmentum and innervate the midbrain ventral tegmental area have been proposed to play a key role in intracranial self-stimulation reward. This mesopontine area also contains GABA neurons. Detailed information is still lacking, however, about the relationship of cholinergic and GABAergic neurons in this region to self-stimulation reward. Therefore, using double immunostaining for Fos as a marker of neuronal activity and choline acetyltransferase as a marker of cholinergic neurons, or for Fos and GABA, we investigated whether self-stimulation of the medial forebrain bundle induces Fos expression within cholinergic and GABAergic neurons in two regions of the mesopontine tegmentum, i.e., pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus. Self-stimulation of the medial forebrain bundle for 1 h induced a large increase in the number of cells expressing Fos in both the pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus, when compared to control brains. However, the self-stimulation-induced expression of Fos was restricted mostly to GABA-, but not choline acetyltransferase-, immunostained cells. We also examined, using microdialysis, whether self-stimulation increases acetylcholine efflux in the ventral tegmental area, a terminal region of the mesopontine tegmentum cholinergic pathway. One hour of self-stimulation significantly increased acetylcholine efflux from this terminal area. These results indicate that intracranial self-stimulation of the medial forebrain bundle may increase acetylcholine release without affecting expression of Fos in cholinergic neurons, while the same stimulation may induce Fos expression in GABAergic neurons of the mesopontine tegmentum. GABAergic as well as cholinergic neurons in this area appear to be activated by self-stimulation reward in the medial forebrain bundle.

Effects of physostigmine and calcium on acetylcholine efflux from the hippocampus of freely moving rats as determined by in vivo microdialysis and a radioimmunoassay

The effects varying the concentration of Ca 2+ in perfused artificial cerebrospinal fluid ([Ca 2+] csf) on basal acetylcholine (ACh) efflux from the hippocampus of freely moving rats, in the presence and absence of the cholinesterase (ChE) inhibitor physostigmine, were investigated using in vivo microdialysis and a highly specific radioimmunoassay for ACh. In the absence of physostigmine, basal ACh efflux was 3.4±0.7 pg/30 min (mean±SEM) at [Ca 2+] csf=1.26 mM. Stepwise increases in [Ca 2+] csf elicited a gradual increase in ACh efflux that was significant at [Ca 2+] csf=5.04 mM. Inhibition of ChE by addition of 10 μM physostigmine to the perfusate increased the efflux of ACh to 103.2±21.1 pg/30 min ([Ca 2+] csf=1.26 mM), and the efflux was augmented still further by increasing [Ca 2+] csf, a change that became significant at [Ca 2+] csf=3.78. These results illustrate the sensitivity of basal ACh efflux from the hippocampus to changes in the extracellular Ca 2+ concentration, and suggest that a more accurate picture of hippocampal cholinergic activity is obtained by microdialysis using normal artificial cerebrospinal fluid, under physiological conditions, rather than in the presence of a ChE inhibitor.

Age-related attenuation of stimulated cortical acetylcholine release in basal forebrain-lesioned rats

In vivo microdialysis was used to measure the effects of partial deafferentation of cortical cholinergic inputs on acetylcholine efflux in young (four to seven months) and aged (24–28 months) male F344/BNNIA rats. Partial deafferentation was produced by bilateral infusions of the immunotoxin 192 immunoglobulin G-saporin (0.56 μg/1.0 μl) or its vehicle solution into the ventral pallidum/substantia innominata region of the basal forebrain. The lesion produced comparable (65%) decreases in basal cortical acetylcholine efflux in young and aged rats. Presentation of a complex environmental stimulus (exposure to darkness/palatable food), in conjunction with the systemic administration of the benzodiazepine receptor weak inverse agonist ZK 93 426, increased cortical acetylcholine efflux in young shams, aged shams and young lesioned rats, but not in aged lesioned rats. Administration of the benzodiazepine receptor partial inverse agonist FG 7142, in the absence of the environmental stimulus, comparably stimulated cortical acetylcholine efflux in young and aged sham rats. FG 7142-induced increases in acetylcholine efflux were attenuated by approximately 50% following partial deafferentation in both young and aged rats. These results suggests that, under certain conditions, ageing potently interacts with the integrity of the cortical cholinergic afferent system. The effects of ageing on cortical cholinergic function may be most potently revealed by experiments assessing age-related limitations in the responsiveness of a partially deafferented cholinergic system to certain behavioral and/or pharmacological stimuli.

Effects of local cholinesterase inhibition on acetylcholine release assessed simultaneously in prefrontal and frontoparietal cortex

To investigate whether acetylcholine is released in a similar fashion in different regions of the cortex, in vivo microdialysis was used to measure acetylcholine efflux simultaneously in the medial prefrontal and the frontoparietal cortex, under both basal conditions and following tactile stimulation. Additionally, the effects of including two different concentrations (0.05 μM and 0.5 μM) of a cholinesterase inhibitor (neostigmine) in the perfusion fluid were assessed. Basal levels of acetylcholine (i.e. during non-stimulated sessions) were similar in medial prefrontal and frontoparietal areas. Tactile stimulation reliably increased acetylcholine efflux in a similar fashion (up to 140% increase above baseline) in both cortical areas studied. Predictably, the higher concentration of neostigmine (0.5 μM) increased basal acetylcholine efflux by about 150% from levels observed with the lower neostigmine concentration (0.05 μM), but the concentration of local neostigmine had no effect on either the magnitude or the duration of the increased acetylcholine efflux following tactile stimulation. These results suggest that the pattern of acetylcholine release may be comparable in different areas of the cortex, supporting the idea that cholinergic projections from the basal forebrain to the cortex represent a globally regulated system. Furthermore, while the inclusion of neostigmine in perfusion fluid must be taken into account when interpreting acetylcholine efflux data, it appears that concentrations of up to 0.5 μM do not interfere fundamentally with the lability of cortical acetylcholine efflux in response to behavioural stimulation.

2116 Effects of naitric oxide on yawning evoked by stimulation of paraventricular nucleus of rats

The cholinergic neurons which originate in the mesopontine tegmentum and innervate the midbrain ventral tegmental area have been proposed to play a key role in intracranial self-stimulation reward. This mesopontine area also contains GABA neurons. Detailed information is still lacking, however, about the relationship of cholinergic and GABAergic neurons in this region to self-stimulation reward. Therefore, using double immunostaining for Fos as a marker of neuronal activity and choline acetyltransferase as a marker of cholinergic neurons, or for Fos and GABA, we investigated whether self-stimulation of the medial forebrain bundle induces Fos expression within cholinergic and GABAergic neurons in two regions of the mesopontine tegmentum, i.e., pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus. Self-stimulation of the medial forebrain bundle for 1 h induced a large increase in the number of cells expressing Fos in both the pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus, when compared to control brains. However, the self-stimulation-induced expression of Fos was restricted mostly to GABA-, but not choline acetyltransferase-, immunostained cells. We also examined, using microdialysis, whether self-stimulation increases acetylcholine efflux in the ventral tegmental area, a terminal region of the mesopontine tegmentum cholinergic pathway. One hour of self-stimulation significantly increased acetylcholine efflux from this terminal area.These results indicate that intracranial self-stimulation of the medial forebrain bundle may increase acetylcholine release without affecting expression of Fos in cholinergic neurons, while the same stimulation may induce Fos expression in GABAergic neurons of the mesopontine tegmentum. GABAergic as well as cholinergic neurons in this area appear to be activated by self-stimulation reward in the medial forebrain bundle.

Acetylcholine output from the ischemic rat cerebral cortex: effects of adenosine agonists

The efflux of acetylcholine (ACh) from the ischemic rat cerebral cortex was examined using the cortical cup technique and an HPLC with electrochemical detection assay. Four vessel occlusion of the cerebral circulation caused a rapid increase in ACh efflux into the cortical superfusates, which was then sustained during the 20 min period of occlusion. Reperfusion was associated with a rapid return of ACh efflux to basal levels. The A 1 and A 2 selective adenosine receptor agonists, N 6-cyclopentyladenosine (10 −8 and 10 −10 M) and CGS 21680 (10 −8), failed to significantly alter ischemia-evoked release of ACh. Because ACh is known to enhance NMDA receptor mediated neuronal depolarization and intracellular Ca 2+ levels, and to potentiate l-glutamate-induced neural degeneration, the present findings suggest that ACh could contribute to ischemic brain injury.

Inhibition of striatal acetylcholine release by endogenous serotonin

We have examined the hypothesis that endogenous serotonin (5-HT) exerts an inhibitory influence on the release of acetylcholine (ACh) in striatum. Striatal slices were prepared from adult rats, preincubated with [ 3H]choline, superfused, and exposed to electrical field stimulation. The stimulation-induced overflow of tritium into the superfusate was used as a measure of ACh release. We observed that fluoxetine, an inhibitor of 5-HT uptake, reduced ACh overflow in slices prepared from caudal striatum, an area of high 5-HT concentration, but not in slices from rostral striatum, an area of low 5-HT concentration. Moreover, methysergide, a 5-HT antagonist, increased ACh efflux in caudal but not rostral striatum. Finally, direct activation of 5-HT receptors with the 5-HT agonist, quipazine, inhibited stimulation-induced ACh overflow in both rostral and caudal striatum. These results suggest that endogenous 5-HT normally is capable of inhibiting striatal ACh release, and that the extent of the modulation is related to the degree of serotonergic innervation. In addition, 5-HT receptors capable of modulating ACh release are present in 5-HT-poor rostral striatum, as well as in 5-HT-rich caudal striatum.