Diabetes affects millions of people worldwide. This devastating disease dramatically increases the risk of developing cardiovascular disorders. A hallmark metabolic abnormality in diabetes is hyperglycemia, which contributes to the pathogenesis of cardiovascular complications. These cardiovascular complications are, at least in part, related to hyperglycemia-induced molecular and cellular changes in the cells making up blood vessels. Whereas the mechanisms mediating endothelial dysfunction during hyperglycemia have been extensively examined, much less is known about how hyperglycemia impacts vascular smooth muscle function. Vascular smooth muscle function is exquisitely regulated by many ion channels, including several members of the potassium (K+) channel superfamily and voltage-gated L-type Ca2+ channels. Modulation of vascular smooth muscle ion channels function by hyperglycemia is emerging as a key contributor to vascular dysfunction in diabetes. In this review, we summarize the current understanding of how diabetic hyperglycemia modulates the activity of these ion channels in vascular smooth muscle. We examine underlying mechanisms, general properties, and physiological relevance in the context of myogenic tone and vascular reactivity.

The L-type Ca2+ channel CaV1.2 is essential for arterial myocyte excitability, gene expression and contraction. Elevations in extracellular glucose (hyperglycemia) potentiate vascular L-type Ca2+ channel via PKA, but the underlying mechanisms are unclear. Here, we find that cAMP synthesis in response to elevated glucose and the selective P2Y11 agonist NF546 is blocked by disruption of A-kinase anchoring protein 5 (AKAP5) function in arterial myocytes. Glucose and NF546-induced potentiation of L-type Ca2+ channels, vasoconstriction and decreased blood flow are prevented in AKAP5 null arterial myocytes/arteries. These responses are nucleated via the AKAP5-dependent clustering of P2Y11/ P2Y11-like receptors, AC5, PKA and CaV1.2 into nanocomplexes at the plasma membrane of human and mouse arterial myocytes. Hence, data reveal an AKAP5 signaling module that regulates L-type Ca2+ channel activity and vascular reactivity upon elevated glucose. This AKAP5-anchored nanocomplex may contribute to vascular complications during diabetic hyperglycemia.

BACKGROUND: L-type Ca V 1.2 channels undergo cooperative gating to regulate cell function, although mechanisms are unclear. This study tests the hypothesis that phosphorylation of the Ca V 1.2 pore-forming subunit α1 C at S1928 mediates vascular Ca V 1.2 cooperativity during diabetic hyperglycemia. METHODS: A multiscale approach including patch-clamp electrophysiology, super-resolution nanoscopy, proximity ligation assay, pressure myography, and Laser Speckle imaging was implemented to examine Ca V 1.2 cooperativity, α1 C clustering, myogenic tone, and blood flow in human and mouse arterial myocytes/vessels. RESULTS: Ca V 1.2 activity and cooperative gating increase in arterial myocytes from patients with type 2 diabetes and type 1 diabetic mice, and in wild-type mouse arterial myocytes after elevating extracellular glucose. These changes were prevented in wild-type cells pre-exposed to a PKA inhibitor or cells from knock-in S1928A but not S1700A mice. In addition, α1 C clustering at the surface membrane of wild-type, but not wild-type cells pre-exposed to PKA or P2Y 11 inhibitors and S1928A arterial myocytes, was elevated upon hyperglycemia and diabetes. Ca V 1.2 spatial and gating remodeling correlated with enhanced arterial myocyte Ca 2+ influx and contractility and in vivo reduction in arterial diameter and blood flow upon hyperglycemia and diabetes in wild-type but not S1928A cells/mice. CONCLUSIONS: These results suggest that PKA-dependent pS1928 promotes the spatial reorganization of vascular α1 C into “superclusters” upon hyperglycemia and diabetes. This triggers Ca V 1.2 activity and cooperativity, directly impacting vascular reactivity. The results may lay the foundation for developing therapeutics to correct Ca V 1.2 and arterial function during diabetic hyperglycemia.

Cigarette smoke, including secondhand smoke (SHS), has significant detrimental vascular effects, but its effects on myogenic tone of small resistance arteries and the underlying mechanisms are understudied. Although it is apparent that SHS contributes to endothelial dysfunction, much less is known about how this toxicant alters arterial myocyte contraction, leading to alterations in myogenic tone. The study's goal is to determine the effects of SHS on mesenteric arterial myocyte contractility and excitability. C57BL/6J male mice were randomly assigned to either filtered air (FA) or SHS (6 hours/day, 5 days/week) exposed groups for a 4, 8, or 12-weeks period. Third and fourth-order mesenteric arteries and arterial myocytes were acutely isolated and evaluated with pressure myography and patch clamp electrophysiology, respectively. Myogenic tone was found to be elevated in mesenteric arteries from mice exposed to SHS for 12 weeks but not for 4 or 8 weeks. These results were correlated with an increase in L-type Ca2+ channel activity in mesenteric arterial myocytes after 12 weeks of SHS exposure. Moreover, 12 weeks SHS exposed arterial myocytes have reduced total potassium channel current density, which correlates with a depolarized membrane potential (Vm). These results suggest that SHS exposure induces alterations in key ionic conductances that modulate arterial myocyte contractility and myogenic tone. Thus, chronic exposure to an environmentally relevant concentration of SHS impairs mesenteric arterial myocyte electrophysiology and myogenic tone, which may contribute to increased blood pressure and risks of developing vascular complications due to passive exposure to cigarette smoke.

The L‐type Ca2+ channel CaV1.2 plays key roles in cell excitability, muscle contraction and gene expression. These channels have been shown to gate in unison (e.g. coupled gating) in several cell types. CaV1.2 coupling amplifies Ca2+ influx, which can regulate cell function. In arterial myocytes, CaV1.2 coupling is increased upon elevations in extracellular glucose (HG) and in cells from diabetic patients and animal models of diabetes. Yet, the mechanisms for induction of this gating modality in CaV1.2 channels are poorly understood. Here, we tested the hypothesis that phosphorylation of CaV1.2 at S1928 is critical for induction of CaV1.2 clustering, which contributes to CaV1.2 coupling, amplification of Ca2+ influx and arterial myocyte contraction in response to HG and diabetes. Using a multiscale approach, we found that HG induced an increase in CaV1.2‐mediated activity, open probability, and coupling frequency and strength in WT arterial myocytes, but not cells from a mouse in which S1928 was mutated to alanine to prevent the phosphorylation of this site (e.g. S1928A). Wild type‐like CaV1.2 response to HG was observed in arterial myocytes from a mouse in which S1700 (another key CaV1.2 phosphorylation site) was prevented (e.g. S1700A), thus underscoring the importance S1928 phosphorylation on CaV1.2 function. Super‐resolution nanoscopy and proximity ligation assay (PLA) revealed an increase in CaV1.2 clustering at the surface membrane of WT, but not S1928A arterial myocytes after acute HG incubation. Similar HG‐induced CaV1.2 biophysical and structural alterations were recapitulated in arterial myocytes from WT diabetic mice (streptozotocin model) and human with diabetes, and these changes were completely absent in cells from diabetic S1928A mice. These results suggest a key role for S1928 phosphorylation in modulating CaV1.2 spatial distribution and gating mode upon HG and diabetes. We propose that our work may lay the foundation for novel therapeutic strategies with single amino acid accuracy to correct channel function and vascular reactivity during pathological conditions.