Biochemical characterization of a filtered synaptoneurosome preparation from guinea pig cerebral cortex: Cyclic adenosine 3':5'-monophosphate-generating systems, receptors, and enzymes

Abstract

A particulate preparation was obtained by low speed centrifugation of guinea pig cerebral cortical homogenates prepared with a Krebs-Henseleit buffer. Light microscopic examination, using a reflected light differential interference contrast system, reveals the presence of intact neurons, axonal fragments, glial cells, and erythrocytes along with an abundance of small spherical entities (diameter about 1.1 μm) and snowman-shaped entities (diameter of larger sphere about 1.1 μm, diameter of attached smaller sphere about 0.6 μm). Many unattached smaller spherical entities are also present (diameter about 0.6 μm). Pressure filtration through 5- or 10-μm Millipore filters, followed by low speed centrifugation and resuspension, removes most of the larger entities to afford a suspension composed mainly of the small spherical and snowman-shaped entities. Electron microscopic examination reveals the presence of many synaptosomes with attached resealed postsynaptic entities. It is proposed that these correspond to the snowman-shaped entities to be termed synaptoneurosomes. Accumulations of cyclic AMP elicited by 2-chloroadenosine and histamine, and by combinations of 2-chloroadenosine, histamine, norepinephrine, and forskolin, are lower in filtered than in unfiltered preparations, whereas accumulations elicited by forskolin are unchanged. Levels of adenylate cyclase are reduced by filtration, whereas levels of phosphodiesterase are unchanged. Filtration reduces levels of markers for whole cells and endothelial cells, whereas neuronal markers such as acetylcholinesterase activity and norepinephrine uptake are increased. Levels of S-100 protein, a marker for glial cells, are not significantly decreased. There is no apparent change in the density of many receptors or ion channels. Levels of A1-adenosine and H1-histamine receptors are increased, whereas levels of so-called peripheral benzodiazepine-binding sites are decreased.