Greenwood, I. A. | bioRxiv Science

bioRxiv2025

Identification of sodium/myo-inositol transporter 1 as a major determinant of arterial contractility

BackgroundAs the sodium/ myoinositol transporter (SMIT1) is a positive regulator of Kv7.4/7.5 channels in arterial smooth muscle we postulated that altering SMIT1 expression could have a major impact upon vascular reactivity. Consequently, this study aimed to characterise the effects of changes of SMIT1 membrane abundance on vascular tone and the molecular mechanisms involved. MethodsIsometric tension recording on 2nd order mesenteric arteries and left anterior coronary arteries from male and female Wistar Han rats. Whole artery membrane potential recording. Single cell and whole artery antibody-based imaging. Morpholino based protein knockdown of SMIT1. ResultsMorpholino-mediated knockdown of SMIT1 enhanced U46619- and methoxamine-mediated contractions of mesenteric artery whilst impairing relaxations to the Kv7 activator ML213, isoprenaline and CGRP. Conversely, augmenting SMIT1 membrane abundance by raising external osmolarity with 150 mM raffinose, impaired receptor-mediated contractions of mesenteric and coronary arteries, augmented relaxations to ML213 and adenosine as well as producing membrane potential hyperpolarisation. Proximity ligation assays revealed that raffinose incubation increased the association of SMIT-Kv7.4/7.5 as well as Kv7.4 and G{beta}{gamma} subunits. The SGK1 inhibitor EMD638683 prevented the raffinose-induced increase in SMIT1 and the anti-contractile effect. ConclusionToggling the membrane abundance of SMIT1 had a dramatic effect on the arterial response to vasonstrictors and vasodilators mediated by greater coordination of Kv7 channels and G{beta}{gamma} subunits. This work identified SMT1-Kv7 channel complexes and SGK1 regulation as key modulators of arterial responsiveness.

cell biology↗

bioRxiv2023

Crucial role for Sodium Hydrogen Exchangers in SGLT2 inhibitor-induced arterial relaxations

1IntroductionSodium dependent glucose transporter 2 (SGLT2 or SLC5A2) inhibitors effectively lower blood glucose and are also approved treatments for heart failure independent of raised glucose. One component of the cardioprotective effect is reduced cardiac afterload but the mechanisms underlying peripheral relaxation are ill defined and variable. We speculated that SGLT2 inhibitors promoted arterial relaxation via the release of the potent vasodilator calcitonin gene-related peptide (CGRP) from sensory nerves independent of glucose transport. Experimental approachThe functional effects of SGLT2 inhibitors (dapagliflozin, empagliflozin, ertugliflozin) and the sodium/hydrogen exchanger 1 (NHE1) blocker cariporide were determined on pre-contracted mesenteric and renal arteries from male Wistar rats using Wire-Myography. SGLT2, NHE1, CGRP and TRPV1 expression in both arteries was determined by Western blot and immunohistochemistry. Kv7.4/5/KCNE4 and TRPV1 currents were measured in the presence and absence of dapagliflozin and empagliflozin. ResultsAll SGLT2 inhibitors produced a concentration dependent relaxation (1{micro}M-100{micro}M) of mesenteric arteries that was considerably greater than in renal arteries. Cariporide relaxed mesenteric arteries but not renal arteries. Immunohistochemistry with TRPV1 and CGRP antibodies revealed a dense innervation of sensory nerves in mesenteric arteries that was absent in renal arteries. Consistent with a greater sensory nerve component, the TRPV1 agonist capsaicin produced significantly greater relaxations in mesenteric arteries compared to renal arteries. Relaxations to dapagliflozin, empagliflozin and cariporide were attenuated by incubation with the CGRP receptor antagonist BIBN-4096, the Kv7 blocker linopirdine and the TRPV1 antagonist AMG-517 as well as by depletion of neuronal CGRP. Neither dapagliflozin nor empagliflozin directly activated heterologously expressed TRPV1 channels or Kv7 channels. Strikingly, only NHE1 colocalised with TRPV1 in sensory nerves, and cariporide pre-application prevented the relaxant response to SGLT2 inhibitors. ConclusionsSGLT2 inhibitors relax mesenteric arteries by a novel mechanism involving the release of CGRP from sensory nerves following inhibition of the Na+/H+ exchanger.

physiology↗