Phospholipase C enzymes hydrolyze the rare membrane lipids phosphatidylinositol-4-phosphate (PIP) and phosphatidylinositol-4,5-bisphosphate (PIP2) PIP into inositol triphosphate (IP3) and diacylglycerol (DAG). Phospholipase C enzymes of the β subtype (PLC-β) function specifically in the signal transduction pathways regulated by Gαq-coupled seven transmembrane receptors. These signaling pathways link plasma membrane receptor activation to intracellular increases in Ca++ through liberation of IP3, activation of protein kinases C through hydrolytic production of DAG, and regulation of ion channels by reduction of PIP2 levels. Second messengers in this bifurcating signaling pathway are utilized ubiquitously to activate physiologic processes as divergent as prostaglandin production, neutrophil chemotaxis, and smooth contraction among many other effects (1). The PLC-β family is composed of gene products from four genetic loci (PLC-β1, -β2, -β3, and β3) with two splice variants from PLC-β1, a and b, that diverge at the C terminus (1). Most tissues express more than one isoform, with PLC-β1 and PLC-β3 being most ubiquitously expressed (2-4); PLC-β4 being most highly expressed in brain (4), and PLC-β2 (2,3) being more restricted to hematopoietic cells. Functional differences between the isoforms include differential sensitivity to activation by Gαq and Gβγ subunits. All known isoforms are activated by Gαq and increased cytosolic Ca++ with varying affinities (2), but only the PLC-β2 and PLC-β3 isoforms appear to be regulated by Gβγ subunits in a physiological context (5). Our interest is the role of PLC-β in regulation of cardiomyocyte hypertrophy in atrial dilatation leading to atrial fibrillation and in ventricular hypertrophy in heart failure. Mouse models and human disease investigations link activation of PLC-β to atrial fibrillation and cardiomyocyte hypertrophy (6,7). Recent studies also link the prenatal intrauterine environment to susceptibility to heart disease in adult animals (8). We are interested in the linkage between heart development and heart disease. In the process of characterizing alterations in PLC-β isoenzymes in heart disease models, we realized that little to no information existed on expression of PLC-β isoforms in embryonic heart development. Therefore, we sought to compare protein and mRNA expression levels of PLCβ1-4 in mice hearts at different developmental stages, specifically E10.5, E 12.5, neonate and weanling (P21) adult. The mRNA and protein expression levels of PLC-β isoforms were compared. PLC-β1, PLC-β3, and PLC-β4, both protein and mRNA, were expressed in E10.5, E12.5 and adult mouse hearts. The mRNA expression level of PLC-β3 was higher than the combined expression of PLCβ1a and PLCβ1b at every developmental age. PLC-β4 mRNA levels were approximately the same as the combined levels of PLC-β1a and PLC-β1b. We were seeing the same expression pattern of all PLCβs isoforms throughout development in mouse hearts. Previous studies on rat neonatal cardiomyocytes in culture or from excised hearts had similar, if not identical findings. Both PLC-β1 and PLC-β3 proteins were found in rat neonatal cardiomyocytes expressed at approximately the same level (10). Study on rats shown that PLC-β1 and PLC-β3 mRNA and proteins were both expressed in hearts. The mRNA and protein levels of PLC-β3 were expressed higher than PLC-β1 (11), which supports our findings. Protein expression of PLC-β1 and PLC-β3 in rodent hearts found that PLC-β3 was expressed but PLCβ1 was not (12). A study on development of rat hearts from day 3-112 showed that mRNA and PLC-β1 proteins were expressed. Slight alterations of PLC-β1 mRNA and protein expression were observed but PLC-β1 both mRNA and proteins were constantly expressed throughout development (13). In summary, we did not observe large differences in expression patterns of PLC-β isoforms through two stages of embryonic heart development, E10 and E12.5, during which the mouse heart is undergoing major structural rearrangements and development to its final architecture. Nor did we observe significant differences in PLC-β expression patterns between embryonic and adult mouse hearts, or between embryonic and rodent neonatal hearts as reported by others. Thus, future studies of heart development and disease will be informed by our observation that PLC-β isoform expression is relatively unchanged across developmental stages and rodent ages.
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