Aging influences cerebrovascular myogenic reactivity and BK channel function in a sex-specific manner.

AIMS: The myogenic reactivity of the middle cerebral arteries (MCA) protects the brain by altering the diameter in response to changes in lumen pressure. Large conductance potassium (BK) channels are known to regulate the myogenic reactivity, yet, it is not clear how aging alters the myogenic reactivity via the BK channel in males and females. Thus, we hypothesize that age-associated changes in BK channel subunits modulate the myogenic reactivity in a sex-specific manner. METHODS AND RESULTS: We used vascular reactivity, patch-clamp, and biochemical methods to measure myogenic reactivity, BK channel function, and expression, respectively in cerebral vessels of adult and aged male and female Sprague Dawley rats. Our results suggest that aging and ovariectomy (OVX) exaggerated the myogenic reactivity of MCA in females but attenuated it in males. Aging induced outward eutrophic remodelling in females but inward hypertrophic remodelling in males. Aging decreased total, Kv, BK channel currents, and spontaneous transient outward currents (STOC) in vascular smooth muscle cells isolated from females, but not in males. Aging increased BKα subunit mRNA and protein both in males and females. However, aging decreased BKß1 subunit protein and mRNA in females only. In males, BKß1 mRNA is increased, but protein is decreased. Iberiotoxin-induced MCA constriction is lower in aged females but higher in aged males. Activation of BKα (10 µM NS1619) and BKß1 (10 µM S-Equol) subunits failed to increase STOCs and were unable to decrease the myogenic reactivity of MCA in aged female but not in aged male rats. OVX decreased, but chronic supplementation of oestradiol restored BK channel expression and function. CONCLUSION: Overall our results suggest that aging or OVX-associated downregulation of the BKß1 expression and function in females results in exaggerated myogenic reactivity of MCA. However, age-associated increase in BK channel function in males attenuated myogenic reactivity of MCA.

Activation of calcium- and voltage-gated potassium channels of large conductance by leukotriene B4.

Calcium/voltage-gated, large conductance potassium (BK) channels control numerous physiological processes, including myogenic tone. BK channel regulation by direct interaction between lipid and channel protein sites has received increasing attention. Leukotrienes (LTA4, LTB4, LTC4, LTD4, and LTE4) are inflammatory lipid mediators. We performed patch clamp studies in Xenopus oocytes that co-expressed BK channel-forming (cbv1) and accessory ß1 subunits cloned from rat cerebral artery myocytes. Leukotrienes were applied at 0.1 nm-10 µm to either leaflet of cell-free membranes at a wide range of [Ca(2+)]i and voltages. Only LTB4 reversibly increased BK steady-state activity (EC50 = 1 nm; Emax reached at 10 nm), with physiological [Ca(2+)]i and voltages favoring this activation. Homomeric cbv1 or cbv1-ß2 channels were LTB4-resistant. Computational modeling predicted that LTB4 docked onto the cholane steroid-sensing site in the BK ß1 transmembrane domain 2 (TM2). Co-application of LTB4 and cholane steroid did not further increase LTB4-induced activation. LTB4 failed to activate ß1 subunit-containing channels when ß1 carried T169A, A176S, or K179I within the docking site. Co-application of LTB4 with LTA4, LTC4, LTD4, or LTE4 suppressed LTB4-induced activation. Inactive leukotrienes docked onto a portion of the site, probably preventing tight docking of LTB4. In summary, we document the ability of two endogenous lipids from different chemical families to share their site of action on a channel accessory subunit. Thus, cross-talk between leukotrienes and cholane steroids might converge on regulation of smooth muscle contractility via BK ß1. Moreover, the identification of LTB4 as a highly potent ligand for BK channels is critical for the future development of ß1-specific BK channel activators.

Polyunsaturated Fatty Acids Modulate the Association between PIK3CA-KCNMB3 Genetic Variants and Insulin Resistance.

BACKGROUND: Neighboring genes PIK3CA and KCNMB3 are both important for insulin signaling and ß-cell function, but their associations with glucose-related traits are unclear. OBJECTIVE: The objective was to examine associations of PIK3CA-KCNMB3 variants with glucose-related traits and potential interaction with dietary fat. DESIGN: We first investigated genetic associations and their modulation by dietary fat in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) study (n = 820). Nine single-nucleotide polymorphisms (SNPs) were selected for analysis, covering more than 80% of the SNPs in the region. We then sought to replicate the findings in the Boston Puerto Rican Health Study (BPRHS) (n = 844). RESULTS: For KCNMB3 missense mutation rs7645550, meta-analysis indicated that homeostasis model assessment of insulin resistance (HOMA-IR) was significantly lower in minor allele T homozygotes compared with major allele C carriers (pooled P-value = 0.004); for another SNP rs1183319, which is in moderate LD with rs7645550, minor allele G carriers had higher HOMA-IR compared with non-carriers in both populations (pooled P-value = 0.028). In GOLDN, rs7645550 T allele homozygotes had lower HOMA-IR only when dietary n-3: n-6 PUFA ratio was low (≤0.11, P = 0.001), but not when it was high (>0.11, P-interaction = 0.033). Similar interaction was observed between rs1183319 and n-3: n-6 PUFA ratio on HOMA-IR (P-interaction = 0.001) in GOLDN. Variance contribution analyses in GOLDN confirmed the genetic association and gene-diet interaction. In BPRHS, dietary n-3: n-6 PUFA ratio significantly modulated the association between rs1183319 and HbA1c (P-interaction = 0.034). CONCLUSION: PIK3CA-KCNMB3 variants are associated with insulin resistance in populations of different ancestries, and are modified by dietary PUFA.

Hypertension of Kcnmb1-/- is linked to deficient K secretion and aldosteronism.

Mice lacking the beta1-subunit (gene, Kcnmb1; protein, BK-beta1) of the large Ca-activated K channel (BK) are hypertensive. This phenotype is thought to result from diminished BK currents in vascular smooth muscle where BK-beta1 is an ancillary subunit. However, the beta1-subunit is also expressed in the renal connecting tubule (CNT), a segment of the aldosterone-sensitive distal nephron, where it associates with BK and facilitates K secretion. Because of the correlation between certain forms of hypertension and renal defects, particularly in the distal nephron, it was determined whether the hypertension of Kcnmb1(-/-) has a renal origin. We found that Kcnmb1(-/-) are hypertensive, volume expanded, and have reduced urinary K and Na clearances. These conditions are exacerbated when the animals are fed a high K diet (5% K; HK). Supplementing HK-fed Kcnmb1(-/-) with eplerenone (mineralocorticoid receptor antagonist) corrected the fluid imbalance and more than 70% of the hypertension. Finally, plasma [aldo] was elevated in Kcnmb1(-/-) under basal conditions (control diet, 0.6% K) and increased significantly more than wild type when fed the HK diet. We conclude that the majority of the hypertension of Kcnmb1(-/-) is due to aldosteronism, resulting from renal potassium retention and hyperkalemia.