Rapid non-genomic activation of cytosolic cyclic AMP-dependent protein kinase activity and [Ca2+](i) by 17 beta-oestradiol in female rat distal colon

1 In this study, the effect of 17 beta-oestradiol on adenosine 3':5'-cyclic monophosphate (cyclic AMP)dependent protein kinase (PKA) activity was inve stigated. 2 Rapid (within 15 min) activation of basal PKA activity was observed in cy tosolic fractions by 17 beta-oestradiol but not by 17 alpha-oestradiol, pro gesterone or testosterone. This stimulation was abolished by the specific P KA inhibitor PKI but not by the classical oestrogen receptor antagonist tam oxifen. 3 17 beta-Oestradiol did not stimulate basal PKA activity in membrane fract ions or in cytosolic fractions from male rats. 4 The increase in cytosolic PKA activity was indirect as (i) it was inhibit ed by the adenylyl cyclase inhibitor SQ22536, (ii) it was mimicked by forsk olin and (iii) 17 beta-oestradiol did not cause a stimulation of basal PKA activity in either type I or type II commercially available PKA holoenzymes . 5 Protein kinase C delta (PKC delta) was directly activated by 17 beta-oest radiol. The specific PKC inhibitor, bisindolylmaleimide I (GF 109203X), abo lished the 17 beta-oestradiol-induced PKA activation. 6 17 beta-Oestradiol stimulated an increase in free intracellular calcium i on concentration ([Ca2+](i)) in isolated female but not male rat colonic cr ypts. This was inhibited by verapamil, nifedipine and zero extracellular [C a2+] but unaffected by tamoxifen. 17 alpha-Oestradiol, testosterone and pro gesterone failed to increase [Ca2+](i). 7 PKC and PKA inhibitors abolished the 17 beta-oestradiol-induced increase in [Ca2+](i). 8 These results demonstrate the existence of a novel 17 beta-oestradiol-spe cific PKA and Ca2+ signalling pathway, which is both sex steroid- and gende r-specific, in rat distal colonic epithelium.