Ultrastructural and morphometric features of the acetylcholine innervation in adult rat parietal cortex: An electron microscopic study in serial sections

Abstract

This study was aimed at characterizing the ultrastructural morphology of the normal acetylcholine (ACh) innervation in adult rat parietal cortex. After immunostaining with a monoclonal antibody against purified rat brain choline acetyltransferase (ChAT), more than 100 immunoreactive axonal varicosities (terminals) from each layer of the Par 1 area were photographed and examined in serial thin sections across their entire volume. These varicosities were relatively small, averaging 0.6 micron in diameter, 1.6 microns 2 in surface, and 0.12 micron 3 in volume. In every layer, a relatively low proportion exhibited a synaptic membrane differentiation (10% in layer I, 14% in II-III, 11% in IV, 21% in V, 14% in VI), for a I-VI average of 14%. These synaptic junctions were usually single, symmetrical (> 99%), and occupied a small portion of the surface of varicosities (< 3%). A majority were found on dendritic branches (76%), some on spines (24%), and none on cell bodies. On the whole, the ACh junctional varicosities were significantly larger than their nonjunctional counterparts, and both synaptic and nonsynaptic varicosities could be observed on the same fiber. A subsample of randomized single thin sections from these whole varicosities yielded similar values for size and synaptic frequency as the result of a stereological extrapolation. Also analyzed in single sections, the microenvironment of the ChAT-immunostained varicosities appeared markedly different from that of unlabeled varicosity profiles randomly selected from their vicinity, mainly due to a lower incidence of synaptically targeted dendritic spines. Thus, the normal ACh innervation of adult rat parietal cortex is predominantly nonjunctional (> 85% of its varicosities), and the composition of the microenvironment of its varicosities suggests some randomness in their distribution at the microscopic level. It is unlikely that these ultrastructural characteristics are exclusive to the parietal region. Among other functional implications, they suggest that this system depends predominantly on volume transmission to exert its modulatory effects on cortical activity.

Simultaneous Release of Glutamate and Acetylcholine from Single Magnocellular “Cholinergic” Basal Forebrain Neurons

“…However, the ability to generate fast EPSCs through corelease of glutamate would allow for additional, rapid, and more punctate effects, more closely geared to the discharge patterns of the BF neurons. Furthermore, because much of the cholinergic innervation is onto dendritic shafts as opposed to pyramidal cell bodies (Beaulieu and Somogyi, 1991;Umbriaco et al, 1994;Mrzljak et al, 1995), it is likely that ACh also modulates the active conductance properties of these dendrites to facilitate or attenuate throughput and summation of synaptic inputs from other sources. If some of these afferents also release glutamate, then this could provide an additional excitatory input with much faster onset and offset rates than that mediated by mAChRs.…”

Selective 5-HT1A Antagonists WAY 100635 and NAD-299 Attenuate the Impairment of Passive Avoidance Caused by Scopolamine in the Rat

“…However, such a mechanism might be counterproductive since AChE inhibitors can also affect 'noncholinergic' neurons via the mode of extrasynaptic transmission (Mrzljak et al, 1998, Umbriaco et al, 1994. In addition, at higher doses AChE inhibitors might 'overstimulate' ACh receptor systems in the brain.…”

Cholinergic Control in Developing Prefrontal–Hippocampal Networks

“…and perfused transcardially with 4% paraformaldehyde dissolved in 0.1 M phosphate buffer, pH 7.4. The brains were removed and postfixed in the same solution for 24 h. Blocks of tissue containing the PFC, cholinergic nuclei, or Hipp were sectioned in either the coronal or sagittal plane at 20 or 50 m. Series of two to four consecutive sections were incubated with the mouse monoclonal antibody against purified rat brain choline acetyltransferase (ChAT) (gift from Dr. C. Cozzari, University of Rome, Italy and Dr. B. K. Hartman, University of Minnesota, Minneapolis, MN), guinea pig polyclonal antibody against recombinant GST-tagged vGluT2 (Millipore), guinea pig polyclonal antibody against rat synthetic linear vGluT1 peptide (Millipore), rabbit polyclonal antibody against recombinant mouse calbindin (CB) (Millipore), or rabbit polyclonal antibody against muscle parvalbumin (PV) (Swant) as described previously (Umbriaco et al, 1994;Mechawar et al, 2000;Henny and Jones, 2008). Briefly, at room temperature (RT), free-floating sections were rinsed in PBS (0.1 M, pH 7.4) and preincubated for 2 h in PBS containing 2% normal horse serum (Vector Laboratories), 1% bovine serum albumin (Roth), and 0.2% Triton X-100 (Roth) to block unspecific staining.…”