Sustained Exposure to the SGLT2 Inhibitor Ertugliflozin, But Not NHE‐1 Inhibition By Cariporide, Attenuates Adrenergic Stimulation of Cytosolic CA2+ Levels in Spontaneously Contracting Cardiac Myocytes

Abstract

IntroductionInhibitors of sodium‐glucose cotransporter 2 (SGLT2i) target renal SGLT2 to reduce reabsorption of glucose, facilitate urinary glucose excretion, and lower blood glucose levels in patients with Type II diabetes (T2DM). The drugs, however, also decrease mortality and heart failure (HF) hospitalizations in T2DM patients. Patients with HF exhibit sympathetic overdrive and cytosolic Ca2+ overload in cardiac myocytes. Because SGLT2i harbor cardioprotective effects, we hypothesize that the SGLT2i ertugliflozin (ERTU) will lower cytosolic Ca2+ concentration ([Ca2+]cyt) in metabolically active and contracting cardiac myocytes after adrenergic stimulation.MethodsERTU was reconstituted in DMSO based on manufacture’s recommendation. Cardiac myocytes were isolated from 0–2‐day old mice and plated on coated (0.2% v/v gelatin, 1 μg/ml fibronectin, and 1 μg/ml laminin) glass cover slips. After two days, spontaneous contractions were observed, and fetal bovine serum content reduced to limit fibroblast overgrowth. By changing the media daily, spontaneously contracting cardiac myocytes were treated for 72 hours with either 1 µM ERTU, 10 µM cariporide (CARI), ERTU and CARI, or vehicle (0.01% v/v DMSO). After 72 hours, cells were loaded with FURA‐2/AM for 1 hour, a steady baseline [Ca2+]cyt was first recorded, then cells were stimulated with phenylephrine (100 µM), and [Ca2+]cyt was recorded for additional 30 minutes.ResultsERTU exposure for 72 hours did not alter baseline [Ca2+]cyt in beating cardiac myocytes when compared to vehicle treated controls but reduced the increase in [Ca2+]cyt after adrenergic stimulation with phenylephrine. In contrast, 72‐hour‐long NHE1 inhibition with CARI increased [Ca2+]cyt at baseline when compared to vehicle and ERTU, and further enhanced the increase in [Ca2+]cyt after phenylephrine stimulation. Interestingly, concomitant administration of ERTU and CARI for 72 hours reduced baseline [Ca2+]cyt and inhibited phenylephrine‐mediated increase in [Ca2+]cyt.ConclusionBecause sustained exposure to the SGLT2i ERTU reduced, but the NHE‐1 inhibitor CARI increased [Ca2+]cyt after acute adrenergic stimulation of metabolically active cardiac myocytes, these data indicate for the first time that ERTU’s beneficial effect on [Ca2+]cyt homeostasis is distinct from NHE1 inhibition. Reduced [Ca2+]cyt levels after sympathetic stimulation may contribute to the cardioprotective effect of SGLT2i. Further research is needed on the molecular mechanisms behind ERTU’s influence on cardiac [Ca2+]cyt.