REGULATION OF UTERINE SMOOTH MUSCLE EXCITABILITY

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Preterm births account for only 10 percent of all deliveries, but are associated with over 80 percent of newborn deaths and more than 95 percent of major newborn morbidity. The lack of understanding of the mechansims regulating uterine contraction has hampered progress towards an effective treatment for this reproductive health problem. Currently used tocolytics have little effect in prolonging gestation, necessitating the investigation of other mechansims of prevention. One potential class of therapeutic targets is potassium channels, due to their ability to potently buffer cell excitation. Electrophysiological reports have shown that myometrial cells contain a variety of K+ channel types that may be potential therapeuric targets, including the large-conductance calcium-activated K+ channel (BKCa channel). Blockage of BKCa channels depolarizes myometrial cells and increases contractile activity while activation of these channels by agonists and beta-adrenergic agents induces potent uterine relaxation. Interestingly, the activation of the BKCa channel by phosphorylating agents, Ca2+, or voltage is dependent on whether it is isolated from non-pregnant or pregnant tissue. Recent evidence illustrates the ability of this channel to undergo alternative splicing in the presence of stress hormones, yielding variants that differ in their sensitivity to intracellular Ca2+ and voltage. Sensitivity to these agents is also dependent on the association of the BKCa channel with its ancillary beta subunit. This suggests that alternative splicing or modulation of beta subunit association may be mechanisms regulating BKCa channel diversity in uterine smooth muscle during gestation. While this channel appears to be an important regulatory component of uterine excitability, its role in modulating myometrial contraction during gestation remains unknown. The specific objective of this proposal is to detemine whether modulation of BKCa channel splice variant expression or beta subunit association correlates to a functional difference in uterine excitability during gestation. Alterations in BKCa channel splice variant expression and function during gestation will be investigated by molecular charactization in combination with electrophysiological and contraction measurements in mice. The specific aims of this proposal are to: 1) compare transcript and protein expression patterns of BKCa channel isoforms in mouse uterine smooth muscle during gestaiton, 2) elucidate BKCa channel beta subunit transcript and protein expression during gestation and detemine whether its assembly with the alpha subunit is modulated during pregnancy, 3) determine the contribution of BKCa channel splice variants to the regulation of uterine smooth muscle contraction during gestation, and 4) characterize the expression of the splice variants of the BKCa channel alpha subunit following stimulation with estrogen and progesterone. Preterm labor is a major health problem, especially given the risks it carries with respect to birth defects and the costs associated with premature delivery. Whether the BKCa channel could be a future target for tocolytic drug therapy is thus a significant question to examine.