Abstract 933: The Alpha-1A Adrenergic Receptor Subtype Is Expressed in Only a Sub-Population of Ventricular Myocytes
Background: Recent studies reveal that alpha-1-adrenergic receptors (ARs) have essential adaptive and protective trophic roles in the heart. Among the three alpha-1-AR molecular subtypes, the A, B and D, the alpha-1A subtype protects myocytes from apoptosis. In the aorta, alpha-1A expression is heterogeneous, but it is unknown if this is true in heart.
Hypothesis: The alpha-1A-AR subtype is not expressed in all myocytes.
Methods: We used an alpha-1A knockout (KO) mouse with a LacZ transgene encoding bacterial beta-galactosidase (beta-gal) replacing exactly the alpha-1A first exon (knocked in), and thus controlled by all endogenous alpha-1A gene regulatory elements. In wild type (WT) mice we measured alpha-1A mRNA normalized to beta-actin by quantitative RT-PCR with intron-spanning primers.
Results: Beta-gal in alpha-1A KO mice was present in the same tissues as alpha-1A mRNA in WT mice, and beta-gal levels and activity increased linearly with LacZ copy number, all indicating that beta-gal was a valid surrogate for alpha-1A expression. Myocardial paraffin sections revealed heterogeneous beta-gal staining throughout the ventricular epicardium and endocardium, and less staining in both atria. In isolated adult ventricular myocytes, beta-gal was present in only a fraction of myocytes. The homozygous alpha-1A KO mouse had 60 ± 3% (n = 4 hearts) myocytes positive for beta-gal, and the heterozygous AKO had 22 ± 2% (n = 3) positive myocytes. WT myocytes had no beta-gal. Right ventricular free wall (RV), septum (S), and left ventricular free wall (LV) had equal numbers of beta-gal positive myocytes. Alpha-1A mRNA in WT mice confirmed equal levels in ventricular regions (RV 0.11 ± 0.1, S 0.11 ± 0.1, LV 0.07 ± 0.03, n=3 hearts), and lower levels in atria (RA 0.03 ± 0.03, LA 0.03 ± 0.01, n=3).
Conclusion: The alpha-1A-AR subtype is expressed in only 60% of myocytes throughout both ventricles, and less in atria. These results challenge the notion that all working ventricular myocytes express all ARs equally, and reveal a critical caveat for functional studies in single isolated myocytes. These data also indicate that the trophic effects of the alpha-1A subtype in heart do not require its presence in all myocytes.