Passive Avoidance Training Induces Enhanced Levels of Immunoreactivity for Muscarinic Acetylcholine Receptor and Coexpressed PKC  and MAP-2 in Rat Cortical Neurons

Abstract

Changes in neocortical immunoreactivity (ir) for muscarinic acetylcholine receptors (mAChRs), protein kinase C gamma (PKC gamma), microtubule-associated protein 2 (MAP-2), and the calcium-binding protein parvalbumin (PARV) induced by the performance of a one-trial passive shock avoidance (PSA) task were studied in young adult male Wistar rats. In experiment I, four groups of animals were formed: three control groups (N, naive; H, habituated but nonshocked; and S, habituated and shocked), and a fully trained group (T, habituated and shocked, followed by a retention trial 24 hr after the footshock). Compared to naive animals, the H, S, and T animals all revealed enhanced cortical ir for mAChRs, PKC gamma, and MAP-2 in discrete subsets of cortical neurons in layers 2, 3, and 5, while no changes were found for PARV. The neurons displaying enhanced levels of ir are of the pyramidal and nonpyramidal cell type and are arranged in a columnar manner. Immunofluorescent double-labeling experiments for mAChR, PKC gamma, and MAP-2 revealed that individual cortical neurons localized within the columns display enhanced ir for all three functionally related proteins. Compared to naive animals, all experimental groups revealed significant increases in the total size of cortical areas showing enhanced ir (H, S, and T over N). A further significant increase is found in animals receiving a footshock over nonshocked animals (S over H, respectively). The retention trial, however, did not induce a further increase (T over S). In some of the animals the patterns appeared to be lateralized, in either the left or right hemisphere. In order to test the role of cholinergic innervation in the induction of enhanced mAChR-ir, unilateral lesions of the nucleus basalis magnocellularis (nbm) were performed in experiment II. Apparently, an intact cholinergic innervation from the nbm is not required for the occurrence of the aforementioned columnar patterns. However, when the enhanced columnar patterns in the sensory areas of the cortex are cholinergically deprived, clear deficits in PSA performance are observed. These results indicate that although ACh is not a prerequisite for the induction of enhanced ir for mAChRs in cortical cells, such neurons demand cholinergic neurotransmission for optimal retention of the shock experience. The alterations in ir for coexpressed mAChR, PKC gamma, and MAP-2 in a discrete subset of cholinoceptive cortical neurons arranged in characteristic patterns most likely represent part of the neuronal substrate involved in functional cortical plasticity related to PSA training.

Non-Selectivity of the Monoclonal Antibody M35 for Subtypes of Muscarinic Acetylcholine Receptors

“…M35 has been used as a tool to dissect the interactions between biochemical and physiological events following mAChR stimulation, and to locate mAChRs in a variety of tissues (Table 2). Various studies describe (a) mAChR distribution at peripheral organs and cell types [5,15,18,26,30,35], (b) mAChR distribution at the cellular and subcellular level in the rat forebrain, [21,23,32,33,36,43,44], (c) the neurochemical nature of the M35-positive neurons by immunofluorescence colocalization studies [14,33,44], (d) changes during development, aging and dementia in mAChR distribution in the brain [8,34,36,40], and (e) experimentally induced plasticity in mAChR distribution [6,22,37,39].…”

“…Previously, pharmacological and anatomical data suggested that it is unlikely that M35 binds to a singe subtype of mAChRs [18,34,35], and Vasudevan and co-workers demonstrated in transfected cell lines that M35 does at least recognize the m3 subtype [41]. Knowledge of subtype selectivity of M35 is a necessary step towards a functional interpretation of the observed plasticity in M35 immunoreactivity [6,22,37,38,39]. The aim of the present study was to determine the mAChR subtype selectivity of M35 employing transfected CHO-K1 cells stably expressing human m1-m5 mAChRs separately [7,10].…”

Neuropsychiatric Illness: A Case for Impaired Neuroplasticity and Possible Quantum Processing Derailment in Microtubules

“…Neurons in the auditory cortex and hippocampus affected by training corresponded with the 15% of neurons containing elevated levels of MAP2, many of which are large pyramidal neurons (Woolf, 1993). Learning-related changes in MAP2, muscarinic receptor, and protein kinase C were similarly noted following avoidance training (Van der Zee et al, 1994). To further uncover the mechanisms by which MAP2 participates in the learning process, the N-terminus of MAP2 was truncated in transgenic mice; this led to contextual memory impairment and reduced ability to bind cAMP-dependent kinase leading to reduced phosphorylation of MAP2 (Khuchua et al, 2003).…”

Muscarinic acetylcholine receptor immunoreactivity in the amygdala—II. Fear-induced plasticity

“…Interestingly, the level of mAChR immunoreactivity in the CEA is not necessarily regulated by the degree of cholinergic innervation, a finding which may add to the observed inverse relation between the degree of cholinergic innervation and mAChR immunoreactivity. Behaviorally induced enhancement of mAChRs in the somatosensory cortex was found to be independent of the presence of cholinergic innervation, 51 and cholinergic denervation of the neocortex did not result in a change in the level of mAChR immunoreactivity of the cortical target neurons 59 (Van der Zee E. A., unpublished observations). These data indicate that the regulation of mAChR protein may also be mediated by noncholinergic neurotransmission, determining the level of cholinoceptivity of the CEA neurons.…”