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Developmental changes in contractility and sarcomeric proteins from the early embryonic to the adult stage in the mouse heart

Sharon Siedner, Martina Krüger, Mechthild Schroeter, Doris Metzler, Wilhelm Roell*, Bernd K. Fleischmann*, Juergen Hescheler*, Gabriele Pfitzer and Robert Stehle

Developmental changes in force-generating capacity and Ca²⁺ sensitivity of contraction in murine hearts were correlated with changes in myosin heavy chain (MHC) and troponin (Tn) isoform expression, using Triton-skinned fibres. The maximum Ca²⁺-activated isometric force normalized to the cross-sectional area (F_CSA) increased mainly during embryogenesis and continued to increase at a slower rate until adulthood. During prenatal development, F_CSA increased about 5-fold from embryonic day (E)10.5 to E19.5, while the amount of MHC normalized to the amount of total protein remained constant (from E13.5 to E19.5). This suggests that the development of structural organization of the myofilaments during the embryonic and the fetal period may play an important role for the improvement of force generation. There was an overall decrease of 0.5 pCa units in the Ca²⁺ sensitivity of force generation from E13.5 to the adult, of which the main decrease (0.3 pCa units) occurred within a short time interval, between E19.5 and 7 days after birth (7 days pn). Densitometric analysis of SDS-PAGE and Western blots revealed that the major switches between troponin T (TnT) isoforms occur before E16.5, whereas the transition points of slow skeletal troponin I (ssTnI) to cardiac TnI (cTnI) and of -MHC to -MHC both occur around birth, in temporal correlation with the main decrease in Ca²⁺ sensitivity. To test whether the changes in Ca²⁺ sensitivity are solely based on Tn, the native Tn complex was replaced in fibres from E19.5 and adult hearts with fast skeletal Tn complex (fsTn) purified from rabbit skeletal muscle. The difference in pre-replacement values of pCa₅₀ (-log([Ca²⁺] M^-1)) required for half-maximum force development) between E19.5 (6.05 ± 0.01) and adult fibres (5.64 ± 0.04) was fully abolished after replacement with the exogenous skeletal Tn complex (pCa₅₀ = 6.12 ± 0.05 for both stages). This suggests that the major developmental changes in Ca²⁺ sensitivity of skinned murine myocardium originate primarily from the switch of ssTnI to cTnI.

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