Loss- and gain-of-function mutations in the Type 1 IP3 receptor (IP3R1) affect myogenic constriction of murine cerebral resistance arteries

Abstract

Myogenic reactivity in resistance arteries/arterioles in response to changing intraluminal pressure is a key physiological mechanism for the regulation of blood pressure and flow. Although IP3-mediated intracellular Ca2+ release is known to contribute to the myogenic response, a lack of selective pharmacological agents to target the predominant IP3R1 Ca2+ release channel has prevented detailed investigation of this key mechanism. Mutations in the IP3R1 known to produce either loss-of-function (LOF) or gain-of-function (GOF) phenotypes lead to cerebellar ataxia in humans, yet the effect of these mutations on the cardiovascular system remains unclear. Here, we employed a novel genetic knock-in mouse model with a single point mutation in the S6 helix of IP3R1 that confers either a LOF (D2594A) or GOF (D2594K) phenotype. In endothelium-denuded posterior cerebral arteries of 3-4 month old WT mice, myogenic vasoconstriction was evident at ≥40 mmHg, maximal at ~120 mmHg, and began to decline at >120 mmHg (n=9). In contrast, myogenic constriction was initiated at higher pressures (≥50 mmHg) and was less robust in arteries of heterozygous IP3R1 LOF mice (n=11). In GOF arteries (n=10), myogenic tone developed at a lower pressure (~20 mmHg), was maximal at ~80 mmHg, and declined at >90 mmHg. Interestingly, pressure-induced Ca2+ elevations were similar in Fura-2 loaded arteries from WT, LOF and GOF mice. Using high sensitivity western blotting, we observed a similar pressure-dependent increase in phosphorylated myosin regulatory light chain (LC20) and myosin phosphatase targeting subunit (MYPT1) at 10, 40, 80 and 120 mmHg in arteries from WT, LOF and GOF mice (n=5 each). Although phospho-MYPT1-T855 content increased with intraluminal pressure in WT and LOF arteries, it did not change in vessels of GOF mice. Elevation of intraluminal pressure (i.e., from 10 to 60 or 120 mmHg) significantly decreased G-actin content in WT arteries, did not affect G-actin content in LOF vessels, and caused a biphasic effect in GOF arteries. Stabilizing the actin cytoskeleton with 0.1 mM jasplakinolide normalized myogenic reactivity in LOF arteries and left-shifted myogenic responsiveness in WT vessels; a minimal effect was observed in GOF arteries. Both jasplakinolide and the actin depolymerizing agent latrunculin B (2 mM) inhibited myogenic tone development at intraluminal pressures > 40-60 mmHg. Taken together, these data demonstrate that genetic modification of IP3R1-mediated Ca2+ mobilization affects the myogenic response of cerebral arteries to intraluminal pressure, possibly by affecting the dynamics of the actin cytoskeleton. Functionally, the observed vascular impairments may contribute to neurological dysfunction in patients with LOF and GOF IP3R1 mutations. Funding has been provided by the Canadian Institutes of Health Research: MOP-97988 (WCC), MOP-142467 (APB), PJT-173352 (SRWC) and PJT-155963 (APB, WCC, GRG and SRWC). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.