ABSTRACT
Adenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) and adenosine 5',5'''-P1,P5-pentaphosphate (Ap5A) are stored in and released from rat brain synaptic terminals. In the present study we investigated the hydrolysis of dinucleotides (Ap4A and Ap5A) in synaptosomes from the cerebral cortex of adult rats. Ap4A and Ap5A, but not Ap3A, were hydrolyzed at pH 7.5 in the presence of 20 mM Tris/HCl, 2.0 mM MgCl2, 10 mM glucose and 225 mM sucrose at 37oC. The disappearance of the substrates measured by FPLC on a mono-Q HR column was both time and protein dependent. Since synaptosome integrity was at least 90 percent at the end of the assay, hydrolysis probably occurred by the action of an ecto-enzyme. Extracellular actions of adenine dinucleotides at central nervous system terminate due to the existence of ecto-nucleotidases which specifically cleave these dinucleotides. These enzymes in association with an ATP diphosphohydrolase and a 5'-nucleotidase are able to promote the complete hydrolysis of dinucleotides to adenosine in the synaptic cleft
Subject(s)
Male , Animals , Rats , Acid Anhydride Hydrolases/analysis , Adenosine Triphosphate/metabolism , Cerebral Cortex/enzymology , Dinucleoside Phosphates/metabolism , Synaptosomes/enzymology , Acid Anhydride Hydrolases/physiology , Adenosine Triphosphate/analysis , Cerebral Cortex/chemistry , Chromatography, High Pressure Liquid , Rats, Wistar , Synaptosomes/chemistryABSTRACT
the subcellular localization in brain of an unconventional, calmodulin-binding myosin (myosin-V) found in neurons, astrocytes and other secretory cells of vertebrates has been investigated by probing Western blots of synaptic fractions from rat cerebral cortex with affinity-purified polyclonal antibodies against myosin-V. Myosin-V was detected in intact synaptosomes and in lysed synaptosomes associated with a particulate fration. Our data suggest a role for brain myosin-V in membrane-cytoskeleton function in the synaptic region
Subject(s)
Animals , Rats , Cerebral Cortex/chemistry , Nerve Tissue Proteins/chemistry , Synaptosomes/chemistry , Blotting, WesternABSTRACT
In studies conducted in patients undergoing cardiac catheterizations, some hemodynamic changes were observed after the acute sublingual administration of the angiotensin converting enzyme inhibitors (ACEI) captopril, enalapril, and lisinopril. These changes consisted of an increase in pulmonary artery pressure, pulmonary vascular resistance (PVR) and induction of hypoxia. The pressure changes were transitory and disappeared after 25 min. The possible mechanisms involved in these changes may relate to interactions of the ACEI with peripheral receptor systems for hormones and neurotransmitters. We have thus undertaken the task of evaluating the potential effect of ACEI on biological receptor molecules. We have begun with studies on muscarinic receptors, and the recently characterized neuropeptide Y (NPY) receptors of endothelial cells. Equilibrium binding assays with 3H-QNB have been conducted for muscarinic receptors using rat brain synaptosomes, due to its expression of multiple muscarinic receptors subtypes. In addition 125BH-NPY binding assays were conducted on intact adrenal medullary endothelial cells. Enalapril and captopril, 10(-7) to 10(-3) M, were not able to produce significant inhibition of either muscarinic or NPY receptor probes. The paradoxical changes elicited by sublingual ACEI seems not to involve interaction with muscarinic or NPY receptors
Subject(s)
Animals , Rats , Angiotensin-Converting Enzyme Inhibitors/pharmacology , Endothelium, Vascular/drug effects , Brain Chemistry , Receptors, Muscarinic/drug effects , Receptors, Neuropeptide Y/drug effects , Synaptosomes/drug effects , Adrenal Medulla/blood supply , Cattle , Cells, Cultured , Endothelium, Vascular/chemistry , Hemodynamics/drug effects , Quinuclidinyl Benzilate/metabolism , Receptors, Muscarinic/metabolism , Synaptosomes/chemistryABSTRACT
Crude mitochondrial fractions of rat hypothalamus, incubated in oxygenated Krebs solution (37ºC), released acetylcholinesterase (AChE) (increased of enzyme activity) into the medium, upon depolarization by 50 mM K+;this effect was absent in Ca²+ free medium. Superfused-synaptosomes obtained from subfractionated crude mitochondrial fractions of rat hypothalamus also released AChE, in a calcium-dependent fashion, when dependent fashion, when depolarized by 50 mM K+ or 50 µM veratridine. The veratridine-releasing effect was antagonized by 2 mM tetrodotoxin. Lactic dehydrogenase activity did not change in the medium with the introduction of depolarizing substances