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Methods for the Isolation, Culture, and Functional Characterization of Sinoatrial Node Myocytes from Adult Mice
Published on: October 23, 2016
Changes in the expression of ion channels, connexins and Ca2+-handling proteins in the sino-atrial node during
Eman Abd Allah1, James O Tellez, Joseph Yanni
1Cardiovascular Medicine, Faculty of Medical and Human Sciences, University of Manchester, Core Technology Facility, 46 Grafton Street, Manchester M13 9NT, UK.
Abstract:
There are important postnatal changes in the sino-atrial node (SAN), the pacemaker of the heart. Compared with the neonate, the adult has a slower intrinsic heart rate and a longer SAN action potential. These changes may be due to differences in ion channel expression. Consequently, we investigated postnatal developmental changes in the expression of ion channels and Ca(2+)-handling proteins in the SAN to see whether this is indeed the case. Using quantitative PCR, in situ hybridization and immunohistochemistry, we investigated the expression of ion channels, Ca(2+)-handling proteins and connexins in the SAN from neonatal (2-7 days of age) and adult (∼6 months of age) New Zealand White rabbits. The spontaneous beating rate of adult SAN preparations was 21% slower than that of neonatal preparations. During postnatal development, quantitative PCR revealed a significant decline in the SAN of the following mRNAs: HCN4 (major isoform responsible for I(f)), Na(V)1.5 (responsible for I(Na)), Ca(V)1.3 (in part responsible for I(Ca,L)) and NCX1 (responsible for inward I(NaCa)). These declines could be responsible for the slowing of the pacemaker during postnatal development. There was a significant decline during development in mRNA for delayed rectifier K(+) channel subunits (K(V)1.5, responsible for I(K,ur), K(V)LQT1 and minK, responsible for I(K,s), and ERG, responsible for I(K,r)) and this could explain the prolongation of the action potential. In situ hybridization confirmed the changes observed by quantitative PCR. In addition, immunohistochemistry revealed hypertrophy of nodal cells during postnatal development. Moreover, there were complex changes in the expression of Ca(2+)-handling proteins with age. In summary, there are significant postnatal changes in the expression of ion channels and Ca(2+)-handling proteins in the SAN that could explain the established changes in heart rate and action potential duration that occur during normal development.
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