Premium
A Method to Measure Simultaneously Cyclic AMP and Inositol Phosphate Accumulation in Rat Brain Slices
Author(s) -
Morin D.,
Zini R.,
QuerolFerrer V.,
Sapena R.,
Tillement J. P.
Publication year - 1991
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1991.tb11400.x
Subject(s) - inositol , inositol phosphate , stimulation , second messenger system , chemistry , agonist , receptor , prazosin , propranolol , phosphate , norepinephrine , sugar phosphates , medicine , endocrinology , biochemistry , biology , antagonist , dopamine
The simultaneous measurement of the accumulation of cyclic AMP and inositol phosphates in rat cerebral cortical slices is described. After stimulation, the separation of cyclic AMP and inositol phosphates was achieved using ion‐exchange chromatography and their concentrations were determined by means of a double‐labeling technique, the substrates adenine and inositol being labeled with 14 C and 3 H, respectively. The recoveries were 70–80% for inositol phosphates and 40–50% for cyclic AMP. To test the applicability of the method, norepinephrine was chosen as an agonist, because it is known to stimulate the production of these two second messengers by interacting with α‐ and β‐adrenergic receptors. This procedure is an improvement over existing methods, because we obtained the simultaneous formation of 3 H‐inositol phosphates and [ 14 C]cyclic AMP in a concentration‐dependent process. EC 50 values were similar for the two, 8.5 ± 3.9 μ M for 3 H‐inositol phosphates and 20.2 ± 6.3 μ M for [ 14 C]cyclic AMP, and close to the values obtained when each process was studied alone. The action of adrenergic antagonists was also tested. Propranolol blocked the norepinephrine stimulation of [ 14 C]cyclic AMP, but did not inhibit the norepinephrine stimulation of 3 H‐inositol phosphates. The opposite results were observed with prazosin. Our results suggest that this method could be a useful tool to examine the interaction between these two receptor‐coupled effectors.