A method for studying synaptosomal release of neurotransmitter candidates, as evaluated by studies on cortical cholecystokinin octapeptide release |
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| Authors: | L J Klaff A M Hudson M Paul R P Millar |
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| Institution: | 1. Endocrine and Diabetes Research Group and Isotope and Immunoassay Laboratory, Department of Medicine, Cape Town, South Africa;2. Department of Chemical Pathology University of Cape Town Medical School, Observatory 7925, Cape Town, South Africa |
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| Abstract: | A method for perfusing rat cortical synaptosomes for studying the regulation of cholecystokinin octapeptide (CCK-8) release has been developed and was found to have advantages over the static incubation system. Synaptosomes isolated from rat cortex were suspended in Biogel P2 columns and perfused with Krebs Ringer Bicarbonate buffer. One hundred mM KCl and 75 microM veratine stimulated CCK-8 release, which was Ca++-dependent. The synaptosomes were functionally viable for at least 135 min of incubation as indicated by multiple 100 mM KCl depolarizations and uptake of (3H)-norepinephrine and (14C)-choline. Dopamine and acetylcholine (10(-6)M) stimulated CCK-8 release while serotonin and norepinephrine were without effect. Approximately 20% of total occluded CCK-8 was released from synaptosomes by 100 mM KCl and degradation of CCK-8 was less than 10%. Perfusion of synaptosomes has several advantages over static incubation systems and allows systematic studies on the role of neurotransmitter in the regulation of neuropeptide secretion. |
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| Keywords: | Membrane depolarization Biogenic amines Neurotransmitter uptake |
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