Vitamin C Deficiency Reduces Muscarinic Receptor Coronary Artery Vasoconstriction and Plasma Tetrahydrobiopterin Concentration in Guinea Pigs

Abstract

Vitamin C (vitC) deficiency is associated with increased cardiovascular disease risk, but its specific interplay with arteriolar function is unclear. This study investigates the effect of vitC deficiency in guinea pigs on plasma biopterin status and the vasomotor responses in coronary arteries exposed to vasoconstrictor/-dilator agents. Dunkin Hartley female guinea pigs (n = 32) were randomized to high (1500 mg/kg diet) or low (0 to 50 mg/kg diet) vitC for 10-12 weeks. At euthanasia, coronary artery segments were dissected and mounted in a wire-myograph. Vasomotor responses to potassium, carbachol, sodium nitroprusside (SNP), U46619, sarafotoxin 6c (S6c) and endothelin-1 (ET-1) were recorded. Plasma vitC and tetrahydrobiopterin were measured by HPLC. Plasma vitC status reflected the diets with deficient animals displaying reduced tetrahydrobiopterin. Vasoconstrictor responses to carbachol were significantly decreased in vitC deficient coronary arteries independent of their general vasoconstrictor/vasodilator capacity (p < 0.001). Moreover, in vitC deficient animals, carbachol-induced vasodilator responses correlated with coronary artery diameter (p < 0.001). Inhibition of cyclooxygenases with indomethacin increased carbachol-induced vasoconstriction, suggesting an augmented carbachol-induced release of vasodilator prostanoids. Atropine abolished carbachol-induced vasomotion, supporting a specific muscarinic receptor effect. Arterial responses to SNP, potassium, S6c, U46619 and ET-1 were unaffected by vitC status. The study shows that vitC deficiency decreases tetrahydrobiopterin concentrations and muscarinic receptor mediated contraction in coronary arteries. This attenuated vasoconstrictor response may be linked to altered production of vasoactive arachidonic acid metabolites and reduced muscarinic receptor expression/signaling.

Keywords: Vitamin C; ascorbic acid; biopterins; vascular responses.

Figure 1

(a) Plasma concentrations of BH₄; (b) plasma BH₂:BH₄-ratio. Means ± SEM, *** p < 0.0001 (n = 8).

Figure 2

Contractile responses in coronary arteries. (a) Contractile responses to 60 mM extracellular potassium; (b) contractile responses to cumulative concentrations of ET-1 and U46610 in coronary arteries from control guinea pigs (green) and vitC deficient (red). Means ± SEM (K⁺, n = 16; ET-1, n = 12; U46619, n = 16).

Figure 3

Scatter plots of coronary artery diameter vs. guinea pig body weight (a) and carbachol-induced vasorelaxation compared with coronary artery diameter (b). Control guinea pigs (green squares) and VitC deficient (red triangles).

Figure 4

Log-concentration-response curves of coronary artery segments from vitC versus control guinea pigs. (a) Vasomotor responses to carbachol in coronary artery segments pre-constricted with 40 mM extracellular potassium; (b) in presence of L-NAME; (c) indomethacin; (d) both L-NAME and indomethacin; (e) atropine. (f) Vasodilator responses to sodium nitroprusside (SNP) in coronary arteries pre-constricted with 40 mM extracellular potassium. Control guinea pigs (green) and VitC deficient (red). Means ± SEM (n = 8–16), * p < 0.05, ** p < 0.01, *** p < 0.001.

Figure 5

Log-concentration-response curves of coronary artery segments after treatment with L-NAME and/or indomethacin. The figures illustrate how the combination of inhibitors in artery segments from the same animal modulates the carbachol-induced vasodilator and constrictor responses. Vasomotor responses to carbachol in artery segments pre-constricted with 40 mM extracellular potassium: in absence versus presence of L-NAME in (a) control guinea pigs; (b) vitC deficient. In absence versus presence of indomethacin in (c) control; and (d) vitC deficient, in absence of L-NAME versus presence both L-NAME and indomethacin in (e) control guinea pigs; (f) VitC deficient, and in absence of indomethacin versus presence both L-NAME and indomethacin in (g) control guinea pigs; (h) VitC deficient. Means ± SEM (n = 8–16), * p < 0.05, ** p < 0.01, *** p < 0.001.