Impaired Dopamine D₁Receptor-mediated Vasorelaxation Of Mesenteric Arteries In Obese Zucker Rats

Objective：Obesity and hypertension are two important components of the metabolic syndrome,often coexist in the same individual, and promote the occurrence and development ofcardiovascular disease[1][2]. The reason about obesity-related hypertention is not fullyunderstood. One of the possible etiologies of the metabolic syndrome is insulin resistance[1][2][3][4]. Insulin stimulated sympathetic nerve system activity[5], vascular smooth muscleproliferation[6]and sodium and water retention[7]and the inhibitory effect on prostacyclinsynthesis[8]have been suggested to be involved in the pathogenesis of obesity-relatedhypertension.Dopamine, a well recognized neurotransmitter in the central nervous system, is also animportant modulator of renal and adrenal function, sodium balance, and blood pressure.Dopamine receptors are classified into two subfamilies: the D1-like receptor subfamilyincludes the D1and the D5receptors, while the D2, D3and D4receptors belong to theD2-like receptor subfamily. Dysfunction of the renal dopaminergic system is implicated in the pathogenesis and/or maintenance of hypertension[1][2]. The obese Zucker rat is a modelof metabolic syndrome characterized by hyperinsulinemia, hyperglycemia, andhypertension. Dopamine has been implicated in the development of obesity partly byincreasing food intake due to decreased dopaminergic function, specifically the D2receptor,in the central nervous system[3][4]. D1receptor mediated-natriuresis and diuresis is alsoimpaired in obese Zucker rats[5][6]. Although the kidney is important in the long-termregulation of blood pressure[7], hypertension is also accompanied by increased vascularresistance[8]. Dopamine receptors are expressed in resistance vessels. In rat mesentericartery, D1and D5receptors are expressed in the tunica media while D2-like receptors areexpressed mainly at the endometrial and adventitia-tunica media transitional zone. Theincrease in blood pressure with aging has been related to a decrease in the expression ofarterial D1, D2, and D5receptor[9]. However, the role of the arterial D1receptor in thehypertension of obese Zucker rats is not clear. Our current study tested the hypothesis thatthe vasorelaxant effect of the D1receptor is impaired in obese Zucker rats.Methods:Part I:1、Rats of12-14weeks were used in this experiment. After the tracheotomy wasperformed, the left carotid artery was catheterized with polyethylene-50tubing for bloodpressure monitoring. Plasma glucose concentrations were determined by using Accu-ChekAdvantage glucose monitoring system. Plasma insulin levels were measured via ELISA.2、The vasodilatory response of isolated mesenteric arteries was evaluated using an isometric Mulvany-Halpern small-vessel myograph (model91M610, J.P. Trading, SciencePark, Aarhus, Denmark). The expression and phosphorylation of D1receptors werequantified by immunoblotting and co-immunoprecipitation.Part II:1、 To determine the effect of hyperinsulinemia and hyperglycemia on the function ofthe arterial D1receptor, we studied obese Zucker rats (six to eight-weeks old) fed (6weeks)vehicle or rosiglitazone, an insulin sensitizer (10mg/kg per day); Blood pressure, plasmainsulin and blood glucose and vasodilatory response of isolated mesenteric arteries weremeasurement according to the aforementioned method when rats were12-14weeks old; D1receptor expression and phosphorylation level were compered between administrationgroup and the control group.2、 Lean Zucker rats (eight to ten-weeks old), fed high-fat diet（with50%fat-derivedcalories）to induce hyperinsulinemia or injected intraperitoneally with streptomycin（singleintraperitoneal injection,65mg/kg）to induce hyperglycemia; Blood pressure, plasmainsulin and blood glucose and vasodilatory response of isolated mesenteric arteries weremeasurement according to the aforementioned method when rats were12-14weeks old; D1receptor expression and phosphorylation level were compered between control group andhyperinsulinemia and hyperglycemia groupPart III:In-vitro study, the effects of high insulin and high glucose concentrations on D1receptor expression and phosphorylation were studied in A10cells, a rat thoracicaorta-derived smooth muscle cell line. Since the mitogen-activated protein kinases areimportant downstream signal molecule of insulin, so MAPK inhibitor was added to observethe effect of insulin on D1receptor expression and phosphorylation.Results:Part I:1、 Several variables, including body weight, and fasting plasma glucose and insulinconcentrations and blood pressure, were higher in obese than lean Zucker rats.2、 Fenoldopam, the D1receptor agonist,(10-7M to10-5M) induced a concentration-dependent vasorelaxation in the third-order mesenteric arteries from lean Zucker rats. Theabsence of endothelium did not affect the fenoldopam-induced vasorelaxation, indicating that the fenoldopam-induced vasorelaxation was endothelium-independent.3、 The vasorelaxant effect of fenoldopam was via a D1-like receptor because aD1-like receptor antagonist, SCH23390(10-7M), blocked the fenoldopam-inducedvasorelaxation, although SCH23390, by itself, was without any effect. The impairedvasorelaxant effect of fenoldopam was not a generalized phenomenon, because sodiumnitroprusside (10-10to10-4M)-mediated vasodilation was not impaired in obese Zucker rats.4、 D1receptor expression was lower while basal D1receptor phosphorylation in thirdorder mesenteric arteries was higher in obese than lean Zucker rats.Part II:1、Rosiglitazone reduced plasma insulin and glucose levels and blood pressure of obeseZucker rats. Besides, we found that rosiglitazone restored the impairedfenoldopam-mediated vasorelaxation in obese rats, increased the decreased-expression ofD1receptor, and decreased the increased-phosphorylation of D1receptor in mesentericarteries from obese rats.2、Lean Zucker rats on a high-fat diet for one month developed hyperinsulinemia buthad normal serum glucose and blood pressure. The STZ-treated lean Zucker rats developedslightly high blood pressure and hyperglycemia but with low plasma insulin levels. Bothhyperinsulinemic and hyperglycemic lean Zucker rats had impaired D1receptor-mediatedvasorelaxation, accompanied with decreased D1receptor expression and increased D1receptor phosphorylation, but the dysfunction of the D1receptor was greater in thehyperinsulinemia than hyperglycemia model.Part III:1、 Treatment with insulin or high glucose decreased D1receptor expression andincreased D1receptor phosphorylation in A10cells for24hr.2、 In the presence of a MAP kinase inhibitor, PD98059(10-6M), the effects ofinsulin on D1receptor expression and phosphorylation were partly blocked.Conclusions:In summary, this study shows that both hyperinsulinemia and hyperglycemia impairvascular D1receptor function and these effects were associated with decreased D1receptorexpression and increased D1receptor phosphorylation. Besides, hyperinsulinemia plays amore important role than hyperglycemia in the dysfunction of the arterial D1receptor in obese Zucker rats. Impaired D1receptor-mediated vasorelaxation is involved in thepathogenesis of obesity-related hypertension.