Elastic and Muscular Arteries Differ in Structure, Basal NO Production and Voltage-Gated Ca(2+)-Channels

Abstract

In the last decades, the search for mechanisms underlying progressive arterial stiffening and for interventions to avoid or reverse this process has gained much attention. In general, arterial stiffening displays regional variation and is, for example, during aging more prominent in elastic than in muscular arteries. We hypothesize that besides passive also active regulators of arterial compliance [i.e., endothelial and vascular smooth muscle cell (VSMC) function] differ between these arteries. Hence, it is conceivable that these vessel types will display different time frames of stiffening. To investigate this hypothesis segments of muscular arteries such as femoral and mesenteric arteries and elastic arteries such as the aorta and carotid artery were isolated from female C57Bl6 mice (5-6 months of age, n = 8). Both microscopy and passive stretching of the segments in a myograph confirmed that passive mechanical properties (elastin, collagen) of elastic and muscular arteries were significantly different. Endothelial function, more specifically basal nitric oxide (NO) efficacy, and VSMC function, more specifically L-type voltage-gated Ca(2+) channel (VGCC)-mediated contractions, were determined by α1-adrenoceptor stimulation with phenylephrine (PE) and by gradual depolarization with elevated extracellular K(+) in the absence and presence of eNOS inhibition with L-NAME. PE-mediated isometric contractions significantly increased after inhibition of NO release with L-NAME in elastic, but not in muscular vessel segments. This high basal eNOS activity in elastic vessels was also responsible for shifts of K(+) concentration-contraction curves to higher external K(+). VGCC-mediated contractions were similarly affected by depolarization with elevated K(+) in muscular artery segments or in elastic artery segments in the absence of basal NO. However, K(+)-induced contractions were inhibited by the VGCC blocker diltiazem with significantly higher sensitivity in the muscular arteries, suggestive of different populations of VGCC isoforms in both vessel types. The results from the present study demonstrate that, besides passive arterial wall components, also active functional components contribute to the heterogeneity of arterial compliance along the vascular tree. This crucially facilitates the search for (patho) physiological mechanisms and potential therapeutic targets to treat or reverse large artery stiffening as occurring in aging-induced arterial stiffening.

Keywords: arterial compliance; arterial stiffness; basal nitric oxide; diltiazem; elastic arteries; muscular arteries; voltage-gated calcium channels.

Figure 1

(A) Transverse sections through segments of the aorta, the carotid artery, the mesenteric artery, and the femoral artery from the same mouse. The segments were mounted in the myograph for 8 h and transverse sections were stained with orcein to show the elastin layers in the different blood vessels. (B) Force per lamella as a function of stretch for the aorta (ao, n = 4), the carotid artery (ca, n = 6), the femoral artery (fa, n = 8), and the mesenteric artery (ma, n = 8). (C) Slope of the linear part of the different force-stretch relationships shown in (B). All slopes were significantly (^***P < 0.001) different from each other.

Figure 2

Tension per lamella as a function of time after addition of 10 μM PE in the absence (eNOS active, open circles) and presence (eNOS inhibited, red) of 300 μM L-NAME in the aorta (A), the carotid artery (B), the femoral artery (C), and the mesenteric artery (D). The maximal tension/lamella of aorta (ao), carotid artery (ca), femoral artery (fa), and mesenteric artery (ma) after 600 s tension development by 10 μM PE (E) or 50 mM K⁺ (F) was calculated. White, eNOS active; red, eNOS inhibited; ^*, ^**, ^***P < 0.05, 0.01, 0.001 eNOS inhibited vs. eNOS active.

Figure 3

Relative isometric contractions of the aorta (ao, n = 4), the carotid artery (ca, n = 6), the femoral artery (fa, n = 6), and the mesenteric artery (ma, n = 5) by increasing K⁺ concentrations (depolarization) in the presence and absence of basal NO (A,C, respectively). Basal NO release was inhibited by incubation with 300 μM L-NAME. The first derivative of the individual concentration-response curves of (A,C) are displayed in (B,D) and correlate with window contraction curves because of L-type Ca²⁺ influx (Fransen et al., 2012a,b). The dashed lines correspond with the EC₅₀ of K⁺ or the maximal change of relative tension per concentration of K⁺ with eNOS active, whereas the arrows between (B) and (D) indicate the shifts of the concentration-contraction curves after inhibition of eNOS with L-NAME for aorta (ao), carotid artery (ca), and mesenteric artery (ma). A shift was absent in fa.

Figure 4

Relaxation of 50 mM K⁺-elicited contractions by the VGCC blocker diltiazem in aorta (ao, n = 4), carotid artery (ca, n = 5), femoral artery (fa, n = 6), and mesenteric artery (ma, n = 5). (A). Diltiazem concentration-relaxation curves in the presence of 300 μM L-NAME to inhibit basal NO release (B) Log(IC₅₀) of diltiazem for inhibition of the isometric contractions induced by 50 mM K⁺ in the absence (eNOS active, white) and presence (eNOS inhibited, red) of 300 μM L-NAME to block basal NO release. ^*, ^**, ^***P < 0.05, 0.01, 0.001.