Premium
A non‐capacitative pathway activated by arachidonic acid is the major Ca 2+ entry mechanism in rat A7r5 smooth muscle cells stimulated with low concentrations of vasopressin
Author(s) -
Broad Lisa M.,
Can Toby R.,
Taylor Colin W.
Publication year - 1999
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1999.0121z.x
Subject(s) - protein kinase c , arachidonic acid , phospholipase c , vasopressin , endocrinology , medicine , inositol , cytosol , receptor , vascular smooth muscle , diacylglycerol kinase , protein kinase a , inositol trisphosphate , chemistry , signal transduction , microbiology and biotechnology , biology , kinase , biochemistry , enzyme , smooth muscle
1 Depletion of the Ca 2+ stores of A7r5 cells stimulated Ca 2+ , though not Sr 2+ , entry. Vasopressin (AVP) or platelet‐derived growth factor (PDGF) stimulated Sr 2+ entry. The cells therefore express a capacitative pathway activated by empty stores and a non‐capacitative pathway stimulated by receptors; only the former is permeable to Mn 2+ and only the latter to Sr 2+ . 2 Neither empty stores nor inositol 1,4,5‐trisphosphate (Ins P 3 ) binding to its receptors are required for activation of the non‐capacitative pathway, because microinjection of cells with heparin prevented PDGF‐evoked Ca 2+ mobilization but not Sr 2+ entry. 3 Low concentrations of Gd 3 + irreversibly blocked capacitative Ca 2+ entry without affecting AVP‐evoked Sr 2+ entry. After inhibition of the capacitative pathway with Gd 3 + , AVP evoked a substantial increase in cytosolic [Ca 2+ ], confirming that the non‐capacitative pathway can evoke a significant increase in cytosolic [Ca 2+ ]. 4 Arachidonic acid mimicked the effect of AVP on Sr 2+ entry without stimulating Mn 2+ entry; the Sr 2+ entry was inhibited by 100 μM Gd 3 + , but not by 1 μM Gd 3 + which completely inhibited capacitative Ca 2+ entry. The effects of arachidonic acid did not require its metabolism. 5 AVP‐evoked Sr 2+ entry was unaffected by isotetrandrine, an inhibitor of G protein‐coupled phospholipase A 2 . U73122, an inhibitor of phosphoinositidase C, inhibited AVP‐evoked formation of inositol phosphates and Sr 2+ entry. The effects of phorbol esters and Ro31‐8220 (a protein kinase C inhibitor) established that protein kinase C did not mediate the effects of AVP on the non‐capacitative pathway. An inhibitor of diacylglycerol lipase, RHC‐80267, inhibited AVP‐evoked Sr 2+ entry without affecting capacitative Ca 2+ entry or release of Ca 2+ stores. 6 Selective inhibition of capacitative Ca 2+ entry with Gd 3 + revealed that the non‐capacitative pathway is the major route for the Ca 2+ entry evoked by low AVP concentrations. 7 We conclude that in A7r5 cells, the Ca 2+ entry evoked by low concentrations of AVP is mediated largely by a non‐capacitative pathway directly regulated by arachidonic acid produced by the sequential activities of phosphoinositidase C and diacylglycerol lipase.