Influence of Estrogen on RLC Phosphorylation and Posttetanic Potentiation of Mouse Muscles with and without Skeletal Myosin Light Chain Kinase
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Estrogen may influence myosin regulatory light chain (RLC) phosphorylation and posttetanic potentiation (PTP) in mouse fast twitch muscle; although the signalling pathway for this effect is unknown. The purpose of this study was to test the hypothesis that previously reported estrogen effects on RLC phosphorylation and PTP are mediated via skeletal myosin light chain kinase (skMLCK). To this end, extensor digitorum longus (EDL) muscles from female wildtype and skMLCK deficient (skMLCK-/-) mice were divided into four groups: ovariectomized (OVX) with estrogen (E+), ovariectomized without estrogen (E-), sham surgery (Sham) and intact baseline (BL). At 8 weeks of age, mice in the OVX groups were ovariectomized followed by pellet implantation at 9 weeks of age with either a 0.1 mg of 17β-Estradiol or implantation of a placebo pellet (E+ and E- respectively); sham surgeries were also performed at this time point for both genotypes. Two weeks later EDL muscles were isolated and suspended in vitro (25° C) for determination of RLC phosphorylation and PTP, except for BL groups which began contractile experiments at 9 weeks. Our results showed that RLC phosphorylation measured in muscles frozen immediately after a potentiating stimulus was not different across conditions within either genotype although values for wildtype muscles were significantly (P<0.05) greater than skMLCK-/- muscles. Consistent with these findings, the ratio of concentric twitch force (post PS / pre PS) for wildtype and skMLCK-/- muscles was similar between E+ and E+ groups although values for wildtype were greater than skMLCK-/- muscles (all data P < 0.05). However, we were unable to directly test our hypothesis as a result of unaltered estradiol levels following OVX. The inability to validate estrogen’s beneficial influence on muscle strength and contractibility in this model could be a direct result of interference with further development and growth during estrogen supplementation. Future studies should note the importance of both estrous cycles and further growth of adult mice when working with ovarian hormones. Both of these factors were likely causes of our atypical findings.