Thomas P. Durkin

Memory-improving action of glucose: indirect evidence for a facilitation of hippocampal acetylcholine synthesis

The effect of a 3 g/kg glucose injection on the velocity of the sodium-dependent high-affinity choline uptake mechanism in the hippocampus was both measured in quiet control mice and in mice immediately after training in an operant bar pressing task. Glucose did not significantly change high-affinity choline uptake in resting animals. High-affinity choline uptake in the hippocampus was increased by training in the operant bar pressing task. Glucose significantly reduced the amplitude of the increase in high-affinity choline uptake observed in the trained animals. Similarly, a 3 g/kg glucose injection also attenuated the increase in high-affinity choline uptake observed in animals injected with 1 mg/kg scopolamine. Finally, a 3 g/kg glucose injection significantly attenuated the amnesia produced by a post-training 1 mg/kg scopolamine injection in mice trained for an operant bar pressing task. These results provide additional evidence for an action of glucose on hippocampal cholinergic activity under conditions of high acetylcholine demand. This action may be mediated via an increase in acetyl coenzyme A availability, one of the precursors of acetylcholine. This facilitative effect of glucose on hippocampal acetylcholine synthesis may constitute the physiological basis for its facilitative action on memory and its attenuation of scopolamine amnesia.

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.

On the involvement of the central cholinergic system in memory deficits induced by long term ethanol consumption in mice.

Male mice of the BALB/c strain were given a solution of 12% v/v ethanol as their only source of fluid for 7 months. Memory performance was tested after ethanol was omitted from the diet for 3 to 9 weeks, and was compared with performance of control animals (no ethanol) which had been pair-fed or had received tap water. The spontaneous alternation task that was used consisted of two forced trials (acquisition) followed, at varying intervals ranging from 30 sec to 6 hr, by a free test trial (retention). Experimental subjects exhibited an accelerated rate of decay of spontaneous alternation, reaching chance level at 6 hours. All animals were then tested at this 6-hour interval following injections of either physostigmine or neostigmine that were given before both acquisition and retention (0.05 mg/kg IP). Results showed that physostigmine, but not neostigmine, dramatically improved performance of alcohol-treated subjects. Parallel neurochemical analysis showed that chronic ethanol treatment induced a slight (12%) but significant decrease in hippocampal sodium-dependent high affinity choline uptake. Though these findings suggest that the observed memory deficits (i.e., an accelerated rate of forgetting) might be related to a cholinergic dysfunction, alternative explanations are also proposed.

The durations of hippocampal and cortical cholinergic activation induced by spatial discrimination testing of mice in an eight-arm radial maze decrease as a function of acquisition

Sodium-dependent high-affinity choline uptake velocities in P2 fractions of the hippocampus and cortex of mice were analyzed at different times following both the first (Day 1) and last (Day 9) daily sessions of a spatial discrimination testing procedure in an eight-arm radial maze. Results showed that the immediate (30 s) post-training increase in mean hippocampal and cortical cholinergic activity observed on Day 1 did not significantly vary over days despite a marked and progressive improvement of discrimination performance. In contrast, the duration of these activations was considerably shortened in both structures between Days 1 (more than 1 hr) and 9 (about 15 min). The possible involvement of these changes in memory consolidation processes is discussed.

A comparison of the working memory performances of young and aged mice combined with parallel measures of testing and drug-induced activations of septo-hippocampal and nbm-cortical cholinergic neurones

The spatial working memory performances of young (2 months) and aged (24-26 months) mice of the C57BL/6 strain were compared using a delayed nonmatching to place (DNMTP) protocol in an automated 8-arm radial maze. The aged mice were observed to exhibit a selective and interference-related memory deficit. Parallel neurochemical analysis of the activity of septo-hippocampal and nbm-cortical cholinergic neurones in vivo was conducted using measures of sodium-dependent high-affinity choline uptake. Results showed that whereas the level of cholinergic activity in both brain regions varied less than 10% between young and aged mice in quiet conditions (basal) the activation usually observed at 30-sec posttest (+20-25%) in young mice was greatly attenuated in the frontal cortex and almost totally absent in the hippocampus of aged mice. In view of these results a complementary experiment was carried out in order to test the intrinsic ability of septo-hippocampal cholinergic neurones to activate using acute injection of scopolamine (1 mg/kg IP 20 min) to both young and aged mice in quiet conditions. The drug injection resulted in a very large (+70%) increase in hippocampal high-affinity choline uptake and with amplitudes which did not vary significantly between young and aged subjects. These observations attest to a relatively well-preserved state of central cholinergic neurones and an intact capacity to activate normally when challenged pharmacologically in aged mice. The results strongly suggest that the loss of cholinergic activation and associated memory deficit in aged mice might rather be related to a hypofunction of phasically active transsynaptic processes which normally mediate the activation of these cholinergic pathways during memory testing.

Self-administration of the GABAA antagonist bicuculline into the ventral tegmental area in mice: dependence on D2 dopaminergic mechanisms

Abstract BALB/c mice were unilaterally implanted with a guide cannula, the tip of which was positioned 1.5 mm above the ventral tegmental area (VTA). On each day of the experimental period, a stainless steel injection cannula was inserted into the VTA in order to study the eventual self-administration of a low dose (1.5 ng/50 nl) of bicuculline, a GABAA-antagonist, using a spatial discrimination task in a Y maze. Mice rapidly discriminated between the arm enabling a micro-injection of bicuculline and the neutral arm of the maze, and robust self-administration of this GABAergic antagonist was observed. Once this self-administration response for bicuculline had been fully acquired, the systemic injection of the dopaminergic D2 antagonist sulpiride (50 mg/kg), 30 min before the test, produced a rapid extinction of the self-administration response. Moreover, if this same sulpiride pre-treatment was given during the initial acquisition period mice did not discriminate between the two arms of the Y-maze. These data demonstrate the dopamine D2 dependence of this bicuculline self-administration behavior, and confirm that GABAergic interneurons and/or inputs normally transynaptically inhibit neuronal activity in the mesocorticolimbic dopamine system.

Genotypic variation in the dopaminergic inhibitory control of striatal and hippocampal cholinergic activity in mice

Abstract Genotypic variation in dopaminergic-cholinergic interactions in striatum and hippocampus has been investigated in the C57BL/6 and BALB/c mouse strains. Acute treatment in vivo with increasing doses of haloperidol provided behavioural evidence for significant strain-dependent variation in neuroleptic sensitivity as assessed by catalepsy scores with C57BL/6 mice being the more sensitive strain. Measures of the kinetics of high affinity choline uptake in crude synaptosomal preparations of striatum and hippocampus from haloperidol treated mice indicated parallel strain variations for the dose-related activation (dis-inhibition) of cholinergic activity in these regions. The relative involvements of the D1–D2 receptor subpopulations in these effects was investigated using the D2 selective antagonist sulpiride. Sulpiride was also observed to produce similar strain-dependent activation of cholinergic activity in both brain regions whereas behavioural effects were not marked and catalepsy absent in both strains. Comparative in vitro studies on the sensitivity of the dopamine stimulated adenylate cyclase from striatal membranes of both strains failed to reveal significant differences. Genotypic variation in agonist sensitivity was explored using apomorphine. While BALB/c mice exhibited significantly greater sensitivity than C57BL/6 regarding dose effects on stereotypy and rearing, apomorphine was found to be without significant effect in either strain regarding striatal or hippocampal high affinity choline uptake kinetics. These results constitute additional evidence for the proposal that cholinergic activity in striatum and hippocampus is tonically inhibited by dopaminergic mechanisms mediated by the D2 receptor sub-population and further that their density and/or functional activity exhibits significant genotypic variation.

In vivo modulation of septo-hippocampal cholinergic activity in mice: relationships with spatial reference and working memory performance

Dopaminergic afferents to the septum mediate a tonic and trans-synaptic inhibitory control on the cholinergic neurones of the septo-hippocampal pathway. Lesion of these afferents using 6-hydroxydopamine (6-OHDA) results in a chronic and specific increase of hippocampal cholinergic activity in mice. The consequence of this in vivo modulation of hippocampal cholinergic activity on the acquisition of both a spatial discrimination and a working memory (delayed non-matching to place) task in an 8-arm radial maze by C57BL/6 mice were investigated. Combined neurochemical and behavioural analyses revealed significant correlations between hippocampal sodium-dependent high-affinity choline uptake activation induced by testing and performance measures. In the first experiment 6-OHDA-treated mice compared to control and vehicle-injected mice showed a transient (day 2) but significant facilitation of their spatial discrimination performance which appears to be better related to the working but not to the reference memory component of the task. This hypothesis is strengthened by the results of the second experiment which shows an amelioration of working memory performance when the septo-hippocampal cholinergic pathway is specifically activated in vivo.

GABAergic mediation of indirect transsynaptic control over basal and spatial memory testing-induced activation of septo-hippocampal cholinergic activity in mice *

A neurochemical study of the transsynaptic interactions established between septal GABAergic interneurones and cholinergic septo-hippocampal neurones was conducted using mice. The effects of acute in vivo injections of either muscimol (20-500 ng/0.2 microliter), bicuculline (100 ng-1 micrograms/0.2 microliter) or saline vehicle (0.2 microliter) into the medial septum on septo-hippocampal cholinergic activity were evaluated using measures of hippocampal high affinity choline uptake at 30 min post-injection in two main groups of mice. The first (quiet control) remained in their home cages during the post-injection period whereas the second (active) were submitted, 10 min following injection to a 20-min period of spatial working memory testing in an 8-arm radial maze. Intraseptal injections of either muscimol or bicuculline produced significant (25-50%) inhibition of hippocampal cholinergic activity in quiet conditions (basal) as compared to intact or saline-injected mice. In the active groups, whereas memory testing induced significant cholinergic activation (+15-20%) in intact and saline injected mice at 30 s post-test no significant memory testing-induced activation was observed in either muscimol or bicuculline-injected mice at any dose. The role of septal GABAergic interneurones in the indirect transsynaptic control over the basal and activated states of septo-hippocampal cholinergic activity is discussed with respect to the concept that these complex neuronal interactions contribute to the physiological mechanisms involved in the modulation of working memory performance.

Septal α-noradrenergic antagonism in vivo blocks the testing-induced activation of septo-hippocampal cholinergic neurones and produces a concomitant deficit in working memory performance of mice

In order to test the hypothesis that alpha-noradrenergic receptors in the septum 1) play an important functional role in the mediation of trans-synaptic control of the neurones of the cholinergic septo-hippocampal pathway and 2) produce resultant modulation of working memory performance, we have investigated the effects in vivo of the acute intraseptal injection of an alpha-antagonist, phenoxybenzamine, in mice. Neurochemical analysis was performed using measures of the kinetics of sodium-dependent high-affinity choline uptake in samples of hippocampus from injected mice and their relevant controls in both quiet conditions and immediately following selective working memory testing in an 8-arm radial maze. Results show that whereas the injection of phenoxybenzamine produces no significant alteration of the activity of the cholinergic septo-hippocampal neurones in quiet conditions, the pretrial (20 min) administration of this drug almost totally abolished the usually observed increase in hippocampal cholinergic activity induced by testing. This inhibition of cholinergic activation was associated with a parallel working memory deficit. The results provide further direct support for the hypothesis that septal noradrenergic afferents via alpha-receptors mediate a phasic and net excitatory trans-synaptic influence on the cholinergic septo-hippocampal pathway during working memory testing and thereby significantly contribute to the modulation of the level of working memory performance.