The overall long-term objective of the proposed studies is to understand the molecular mechanisms which control electrical activity in uterine smooth muscle and to identify molecular targets which might be useful in the management of abnormal labor. The apparent induction by estrogen of a myometrial mRNA species which causes expression of a very slow voltage-dependent potassium current in Xenopus oocytes has previously been demonstrated. This mRNA species is detectable at term and proestrus but not at midgestation or metestrus. It was hypothesized that this mRNA species encodes a smooth muscle K+ channel whose level of expression varies during the estrous cycle and pregnancy depending upon estrogen levels. The effect of increasing expression of the slowly activating K+ current may be to enhance membrane excitability by bringing the membrane potential into an optimum range for the operation of voltage-dependent currents necessary for generating spontaneous activity. The main question to be addressed by the proposed studies is: What is the role of changes in the expression of the slowly activating potassium current in the regulation of myometrial excitability by estrogen? First, the sequence encoding the slow K channel will be determined. Full-length cDNA clones for the slow K+ channel will be isolated from a uterine cDNA library and expressed in xenopus oocytes. Second, the physiological regulation of this mRNA species will be studied. The tissue-specific and hormone-regulated expression of the mRNA species encoding the slow K+ channel will be characterized using nucleic acid hybridization methods. Third, the effect of the mRNA regulation on the numbers of K+ channels in the myometrial cells will be examined. Single potassium channels will be studied in acutely dissociated uterine smooth muscle cells using patch techniques. Finally, the effect of suppressing translation of the K channel mRNA on the transformation of electrical properties by estrogen will be examined using hybrid-arrest techniques in myometrial cells treated with estrogen in vitro.
