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M 1 Muscarinic Receptors Stimulate Rapid and Prolonged Phases of Neuronal Nitric Oxide Synthase Activity: Involvement of Different Calcium Pools
Author(s) -
Wotta Diane R.,
Parsons Ann M.,
Hu Jingru,
Grande Andrew W.,
ElFakahany Esam E.
Publication year - 1998
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 - 10.1046/j.1471-4159.1998.71020487.x
Subject(s) - nitric oxide synthase , muscarinic acetylcholine receptor , neuronal nitric oxide synthase , nitric oxide , calcium , receptor , chemistry , neuroscience , muscarinic acetylcholine receptor m1 , endocrinology , medicine , biology , biochemistry
This study shows that activation of M 1 muscarinic receptors, when coexpressed in Chinese hamster ovary (CHO)‐K1 cells with neuronal nitric oxide (NO) synthase (nNOS), produces early and late phases of elevation of both intracellular Ca 2+ concentration and nNOS activity. We examined the relationship between receptor‐mediated increases in intracellular Ca 2+ concentration and activation of nNOS over both short and long intervals using guanosine 3′,5′‐cyclic monophosphate (cGMP) formation as a measure of nNOS activity. The rapid phase of nNOS activation was dependent on release of Ca 2+ from intracellular stores in both the CHO M 1 /nNOS transfected cells and in neuroblastoma (N1E‐115) cells, in which muscarinic receptors and nNOS are endogenously expressed. Two single point mutations in the M 1 muscarinic receptor that have previously been shown to uncouple differentially the receptor from phosphoinositide hydrolysis produced parallel attenuation of the rapid phase of nNOS activation. Characterization of the prolonged phase of nNOS activation was done using the conversion of l ‐[ 3 H]arginine to l ‐[ 3 H]citrulline as well as cGMP formation following stimulation of M 1 muscarinic receptors for 60 min. Both responses were dependent on influx of extracellular Ca 2+ and were accompanied by prolonged formation of NO at functionally effective levels as late as 60 min following receptor activation. Therefore, this study demonstrates for the first time the existence of two mechanistically distinct phases of nNOS activation that are dependent on different sources of Ca 2+ .