The actions of regulatory steroids are manifested in myometrium through altered synthesis of proteins. Protein components of the sarcolemma such as agonist receptors, ion channels and gap junctions are affected. Major changes in excitability and electrocoupling of the smooth muscle cells are the basis for the functional transition from the quiescent organ of pregnancy to the active and expulsive organ of labor. Interference with these changes may provide a means of preventing pre-term labor. Research should be focussed on determining where potential therapeutic agents may act using in vitro methods and animal studies. Our research will identify proteins, the synthesis of which is subject to regulation by estrogens and progestins, that underly some functional changes initiating labor. Creatine Kinase-BB (formerly Induced Protein) will serve as a model protein to test our hypothesis that progesterone serves mainly to tonically constrain synthesis of """"""""activator"""""""" proteins. These proteins are expected to be of two main classes; a) those that are destined to become incorporated into the membrane with roles closely linked to the process of excitation such as agonist rceptors, junctional components and ion channels; and b) enzymes capable of modifying intrinsic sarcolemmal components to render the cell more excitable. Methods are in place to enable in vitro steroid manipulations on surviving explants and myometrial smooth muscle cells have been established in cloned culture. These techniques will permit the exposure of additional sarcolemmal and soluble proteins subject to tight steroidal control by affinity labelling or pulse-labelling of surface and newly synthesized proteins in cloned smooth muscle cells, followed by 2D electrophoresis. Immunoreagents prepared against these steroid regulated proteins will be evaluated for their suitability as probes for studying dynamic changes in the membrane of myometrial cells being exposed to steroidal manipulations. Methods will be developed using the rabbit as a model species because of clear-cut steroidal control of the myometrium; it is planned to extend the work later by applying the methodology to primates with the eventual goal of studying human myometrial cells established into cell culture.
