Involvement of the septohippocampal cholinergic system in representational memory.

To develop an animal model for testing muscarinic agonists, we examined the effects of cholinergic lesions with the ethylcholine aziridinium ion (AF64A) on two types of memory tasks. The tasks provided a distinction between representational and dispositional memory that could be measured in a single paradigm. Young, male Long-Evans rats were trained in a modified T-maze to learn both a discrimination task and a paired-run alternation task. Once animals learned the tasks, they were administered either saline or AF64A (5 nmol into each hippocampus) via stereotaxic technique. One week following surgery, saline-treated animals exhibited comparable performances (P greater than 0.2) on both the discrimination task (90.0 +/- 2.6% correct) and the alternation task (79.5 +/- 5.7%). In contrast, animals treated with AF64A showed a significant impairment of performance (P less than 0.005) on the alternation task (56.1 +/- 1.7%) as compared to the discrimination task (81.6 +/- 5.0%). Performance of the alternation task was significantly lower for AF64A-treated animals than for controls (P less than 0.02). AF64A-treated animals subsequently injected with pilocarpine (1.0 mg/kg, i.p.) showed moderate improvements in performance on the alternation task, while performance on the discrimination task remained unaffected. Immunocytochemical studies of choline acetyltransferase (ChAT) and tyrosine hydroxylase (TH) immunoreactivity indicated a loss of ChAT-positive cells in the septal region in AF64A-injected animals while TH-positive cells in the ventral tegmental area were unaffected by the treatment. The data suggest that AF64A can be used to produce selective lesions of the septohippocampal cholinergic system, which plays a greater role in representational memory than in dispositional memory.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of muscarinic receptors by intrahippocampal injections of gallamine.

Multiple intrahippocampal injections of gallamine impair performance of a representational memory task in rats. The binding of [3H]-(-)-quinuclidinyl benzilate (QNB) to rat brain sections was measured to determine if changes in receptor binding were associated with the deleterious effects of gallamine. [3H]-(-)-QNB binding to sections taken from gallamine-injected animals was compared with binding in saline-injected control animals. Autoradiographic analyses indicated an increase in [3H]-(-)-QNB binding sites within all layers of the cerebral cortex and in the superior colliculus in gallamine-treated animals as compared to saline-injected controls. Significant increases were noted in cortical layers IV and V (P less than 0.025) in gallamine-treated animals. No significant changes (P greater than 0.05) in the number of binding sites were observed in the hippocampus, neostriatum or various thalamic nuclei. The ability of unlabeled pirenzepine, gallamine and carbamylcholine to inhibit 0.2 nM [3H]-(-)-QNB binding also was measured to determine changes in the distribution of receptor subtypes. No significant changes were observed in any brain region for the binding of the selective antagonists pirenzepine and gallamine or the agonist carbamyl-choline. Although other possibilities are considered, the data suggest that an increase in the number of muscarinic receptors may contribute to the observed behavioral deficits associated with long-term gallamine treatment.

Biochemical and behavioral responses of pilocarpine at muscarinic receptor subtypes in the CNS. Comparison with receptor binding and low-energy conformations.

Pilocarpine was tested biochemically in vitro for its ability to stimulate phosphoinositide (PI) turnover in the hippocampus (M1/M3 responses) where it displayed 35% of the maximal carbachol response with an EC50 value of 18 microM, and low-Km GTPase in the cortex (M2 response), where it had 50% of the maximal carbachol response with an EC50 value of 4.5 microM. Behaviorally, pilocarpine was able to restore deficits in a representational memory task (sensitive to M1 antagonists) produced by intrahippocampal injections of AF64A. Twenty-three low-energy conformations of protonated pilocarpine were generated using the program MacroModel. The data indicate that pilocarpine is a partial agonist at both M1 and M2 muscarinic receptors in the CNS. Behaviorally, with respect to the memory task, M1 effects of pilocarpine apparently predominate. It also is conceivable that different conformations of pilocarpine are active as agonists at different muscarinic receptor subtypes.

Evidence for a preferential involvement of M1 muscarinic receptors in representational memory.

The effects of intrahippocampal injections to the M1-selective antagonist pirenzepine and the M2-selective antagonist AF-DX 116 were examined on performance of a representational memory task in rats. Although both antagonists impaired performance, pirenzepine was more potent than AF-DX 116. Pirenzepine (70.8 +/- 2.8% correct) produced a greater deficit than AF-DX 116 (83.3 +/- 0.0%) at 70 micrograms, and the deficit at 10 micrograms (83.3 +/- 2.8%) was equal to that produced by 70 micrograms of AF-DX 116. The data provide additional support for the cholinergic hypothesis of memory and new information regarding the subtypes of muscarinic receptors likely to be involved in representational memory. Based on the greater susceptibility of representational memory to the effects of pirenzepine, it is suggested that M1 receptors in the hippocampus play a greater role in memory function than M2 receptors.