Prostaglandin E2 Modulates Voltage-gated Calcium Channel Expression in Murine Hearts

Prostaglandin E2 (PGE2) acts locally through 4 receptor subtypes (EP1-EP4) in the heart, although EP3 and EP4 predominate in cardiomyocytes. We previously reported that mice with cardiomyocyte-specific overexpression of the EP3 receptor (EP3 TG) exhibit reduced cardiac function and diminished cardiomyocyte contractility. Reduced contractility of cardiomyocytes may result from changes in calcium handling or calcium insensitivity at the myofilament. We therefore hypothesized that PGE2 via EP3 regulates calcium handling proteins to reduce contractility. To test our hypothesis, we performed RNA sequencing on left ventricles of male EP3 TG mice along with their wild type (WT) controls. We found that mRNA of voltage-gated calcium channel (VGCC) beta subunit (CACNb2), Sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA2a) and Ryanodine receptor 2 (RYR2) expression were significantly downregulated in TG mice in addition to significant reductions in Troponin I. Using western blot, significant reduction of CACNb2 protein was found in both male and female left ventricular lysates of EP3 TG compared to age matched EP3 WT (for males WT 1.0± 0.09 vs TG 0.53 ± 0.04, p = 0.0012, for females WT 1.0± 0.03 vs TG 0.53 ± 0.05, p = 0.020) confirming our RNA seq data and suggesting an absence of sexual dimorphism. To validate that downregulation of CACNb2 is specific to cardiomyocytes, we tested the protein expression of CACNb2 in isolated cardiomyocytes from EP3 WT and TG and it was downregulated in TG (WT 1.0± 0.17 vs TG 0.39 ± 0.09, p = 0.0006). Then to test whether CACNb2 is directly regulated by PGE2, we performed time course experiments in which cardiomyocytes were treated with PGE2 and the EP3 specific agonist sulprostone for 30 minutes and 2 hours. CACNb2 was downregulated with PGE2 and sulprostone at both time points (one way-ANOVA with multiple comparisons was used, (F (2,9) = 53.87, P=0.0001). To further test the specificity of EP3 receptor mediated reduction of CACNb2, we transduced adult ventricular cardiomyocytes (AVM) isolated from C57Bl/6 with adenovirus to overexpress EP3 receptor. After transduction, AVM were treated with PGE2 and sulprostone for 2 hours. A significant reduction of CACNb2 protein was seen with EP3 overexpression in vehicle treated AVM compared to vehicle treated GFP control suggesting that CACNb2 downregulation is mediated via EP3 receptor (GFP control 1± 0 vs EP3 overexpressed AVM 0.73 ±0.02, Paired t-test, P=0.0099). In the literature, miR499 has been shown to downregulate CACNb2 by binding to its 3’ untranslated region. To test miR499 regulation by PGE2 and sulprostone, we treated AVM from C57Bl/6 with PGE2 and sulprostone for 30 minutes, 2 hours and 24 hours. PCR was performed to test the expression of miR499 and CACNb2 mRNA. No significant upregulation of miR499 was seen at 30 minutes, 2 hours and 24 hours suggesting that miR499 mediated downregulation is not responsible for PGE2 downregulation of CACNb2. Finally, we conclude that CACNb2 is downregulated by PGE2 through EP3 receptor. Since CACNb2 regulates the pore forming subunit of VGCC (CACNA1c) that controls calcium entry, it suggests that PGE2 plays role in calcium transport in cardiomyocytes. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.