ERK Inhibition Attenuates 5-HT-Induced Contractions in Fetal and Adult Ovine Carotid Arteries

Abstract

Growth and differentiation-related pathways are much more active in immature than in mature, fully differentiated smooth muscle. Because mitogen-activated protein kinases (MAPK) are intimately involved with growth and differentiation, and the extracellular signal-regulated kinase (ERK) subfamily of MAPKs are involved in some contractile responses, the present studies examined the hypothesis that ERKs play an important and age-dependent role in smooth muscle contraction. The MAPK inhibitors PD098059 and UO126 both inhibited serotonin (5-HT) concentration-response relations more effectively in carotid arteries from term fetal lambs, than in corresponding arteries from mature non-pregnant adult sheep. This inhibition involved significant decreases in both the pD2 (adult: 2-fold; fetus: 4- to 15-fold) and the maximum efficacy (adult: 15-19%; fetus: 34-39%) of 5-HT. Accompanying this age-dependent effect on contraction, quantitative Western blot assays revealed that ERK1 and ERK2 abundances were 39% and 164% greater, respectively, in fetal than in adult carotid arteries. The abundance of the putative ERK target, caldesmon, however, was about 7-fold greater in adult than in fetal arteries. Together, the present results support the view that ERK abundance and activity is upregulated in fetal relative to adult arteries, and that one consequence of this upregulation is that the contribution of ERKs to contraction, at least that initiated by 5-HT2a receptors, is greater in fetal than adult carotid arteries. Whereas the phosphorylation mechanisms through which ERKs augment contraction remain uncertain and controversial, the present results suggest that emphasis should be shifted away from caldesmon and toward other critical contractile proteins, and how these proteins may contribute differently to development of agonist-induced contractile force in immature and mature arteries.

Enhanced Transcription of Contractile 5‐Hydroxytryptamine 2A Receptors via Extracellular Signal‐Regulated Kinase 1/2 after Organ Culture of Rat Mesenteric Artery

“…Our results are well in line with the findings that specific ERK1/2 pathway inhibitors PD098059 and UO126 inhibited contractile responses to 5-HT in carotid arteries of sheep and that 5-HT can activate the ERK1/2 pathway in arterial smooth muscle through activation of 5-HT 2A receptors (Xu et al 2002). It is further supported by the fact that 5-HT 2A mediated-contraction is partly dependent on ERK1/2 activation (Banes et al 1999;Pearce et al 2003). Interestingly, one study has revealed the inability of 5-HT to activate JNK and p38 kinase pathways in rat aortic vascular smooth muscle cells (Banes et al 2001).…”

Trophic action of sympathetic nerves reduces arterial smooth muscle Ca2+sensitivity during early post-natal development in rats

“…These proteins mediate the Ca 2+ -independent activation of the contractile machinery and, therefore, represent markers of the Ca 2+ -independent contractile phenotype. While a similar reduction in the ERK1/2 protein level has been reported in developing ovine carotid arteries (Pearce et al 2003), such changes have not yet been described, to the best of our knowledge, for p38 MAPK and RhoK. Notably, inhibition of RhoK activity results in a more prominent depression of contraction in developing saphenous arteries than in arteries from adults rats (Mochalov et al 2008).…”

Multiple activation mechanisms of serotonin-mediated contraction in the carotid arteries obtained from spontaneously hypertensive rats

“…Controversial reports are available regarding 5-HT-mediated signal transduction via MAPKs in vasculature. For instance, 5-HT-mediated contraction is mediated by ERK1/2 activation [10,50,59] and p38 MAPK [20,34,54]. On the other hand, Banes et al [3] found that 5-HT could not activate p38 MAPK and JNK in rat aortic vascular smooth muscle cells.…”

“…2, Table 1). [10,20,50,59]. Pretreatment of carotid arteries with each inhibitor as an MEK/ERK pathway inhibitor PD98059 (10 μmol L −1 ) (Fig.…”

“…HT 5-hydroxytryptamine, NE norepinephrine, SHR spontaneously hypertensive rat, WKY Wistar Kyoto, Emax the maximal response (mN) generated by the constrictors, pD 2 a negative logarithm of EC50 which is the concentration of the agonist producing 50 % of Emax, AUC area under the curve 44 kDa, 1:1000; Cell Signaling Technology #4696), antiphospho-p38 MAPK (43 kDa, 1:1000; Cell Signaling Technology #9211), anti-p38 MAPK (43 kDa, 1:1000; Cell Signaling Technology #9212), anti-phospho-JNK (46 and 54 kDa, 1:1000; Cell Signaling Technology #9251), anti-JNK (46 and 54 kDa, 1:1000; Cell Signaling Technology #9252), anti-phospho-MYPT1 (130 kDa, 1:200; Santa Cruz Biotechnology #sc-17432), anti-MYPT1 (130 kDa, 1:1000; BD Transduction Laboratories #BD612164), anti-phospho-PDK1 (58-68 kDa, 1:1000; Cell Signaling Technology #3061), anti-PDK1 (58-68 kDa, 1:1000; Cell Signaling Technology #3062), anti-ROCK1 (∼160 kDa, 1:500; BD Transduction Laboratories #BD611136), anti-ROCK2 (∼160 kDa, 1:500; BD Transduction Laboratories #BD610623), anti-5HT 2A receptor (53 kDa, 1:500; ImmunoStar #24288), and anti-β-actin (42 kDa, 1:5000; Sigma #A5316).…”