| Summary: | Alterations in the reactivity of blood vessels to neurotransmitters and circulating hormones have been suggested to cause, or contribute to, some of the cardiovascular complications associated with diabetes mellitus. Previous studies have demonstrated that aortae and mesenteric arteries from male rats with streptozotocin (STZ)-induced diabetes of 12 weeks duration are more responsive to the contractile effects of noradrenaline (NA). The increased responsiveness of the diabetic arteries to NA has been reported to result from stimulation of α₁-adrenoceptors. Contraction of vascular smooth muscle induced by NA via α₁-adrenoceptors has been shown to be associated with enhanced metabolism of phosphoinositides. The purpose of the present study was to investigate whether the enhanced contractile responses of aortae and mesenteric arteries from diabetic rats to NA are associated with alterations in phosphoinositide metabolism. Therefore, we compared changes in contractility and phosphoinositide metabolism in response to NA in aortae and mesenteric arteries from male rats with STZ-induced diabetes of 12-14 weeks duration and their age-matched controls. To further investigate the specificity and mechanisms of alterations in these responses, experiments were also conducted using portal veins, and selective α₁-adrenoceptor agonists, potassium chloride and a direct activator of protein kinase C (PKC).
Maximum contractile responses but not sensitivities of aortae and mesenteric arteries from diabetic rats to NA were significantly enhanced compared with responses of arteries from control rats. In contrast,
responses of portal veins from control and diabetic rats to NA were not different from each other. In addition, contractile response of diabetic arteries to KCl were not altered. Contractile responses of both aortae and mesenteric arteries in Ca²⁺-free medium to maximum concentrations of NA, PE and METH were significantly greater in preparations from diabetic than from control rats. Following readdition of Ca²⁺, responses of diabetic arteries to the α-adrenoceptor agonists were also significantly greater than controls. In the absence of extracellular Ca²⁺, contractions of diabetic aortae to a submaximum concentration of NA (10⁻⁶M) were also significantly increased compared with control, while the responses of diabetic mesenteric arteries to this concentration of NA were not significantly different from those of control preparations. Upon readdition of Ca²⁺, contractile responses of both arteries from diabetic rats were significantly greater than those from controls. These data indicate that the maximum contractile responses of diabetic aortae and mesenteric arteries to α₁-adrenoceptor stimulation are associated with increased release of intracellular Ca²⁺. However, α₁-adrenoceptor-mediated Ca²⁺ influx may also be enhanced in arteries from diabetic rats particularly in response to submaximum concentrations of NA.
A maximum concentration of NA (10⁻⁵M) induced a rapid, transient decrease in the level of [³²P]-phosphatidylinositol 4,5-bisphosphate ([³²P]-PtdIns(4,5)P₂), and a time-dependent increase in the formation of [³²P]-phosphatidic acid ([³²P]-PA) and [³H]-inositol phosphates in aortae from both control and diabetic rats. The increase in [³H]-inositol phosphate production was selectively blocked by prazosin. In
both types of aortic preparations, NA also induced a rapid, transient and concentration-related elevation of inositol 1,4,5-trisphosphate (Ins(l,4,5)P₃) content during contraction. The breakdown of [³²P]-PtdIns(4,5)P2, and formation of [³²P]-PA, [³H]-inositol phosphates and Ins(l,4,5)P₃ in response to the maximum concentration of NA were significantly greater in diabetic than in control aortae.
Stimulation of α₁-adrenoceptors by a maximum concentration of NA (3x10⁻⁵M) in mesenteric arteries resulted in a time-dependent increase in the formation of [³H]-inositol phosphates in both control and diabetic rats. NA also caused a rapid and concentration-dependent elevation of Ins(l,4,5)P₃ content in mesenteric arteries from both groups of animals. The increases in [³H]-inositol phosphates and Ins(l,4,5)P₃ levels in diabetic mesenteric arteries in response to the maximum concentration of NA were significantly higher than in control preparations. In contrast, Ins(l,4,5)P₃ production in mesenteric arteries in response to a submaximum concentration of NA (10⁻⁶M) was not significantly different between control and diabetic preparations.
NA also induced a time-dependent increase in the production of [³H]-inositol phosphates in both control and diabetic portal veins. The NA-induced formation of [³H]-inositol phosphates was not significantly different control and diabetic portal veins with a short period of exposure (0.5 min) to the agonist suggesting that the extent of occurrence of early biochemical events including the hydrolysis of PtdIns(4,5)P₂ in response to NA is similar in both types of portal veins.
The role of PKC in mediating enhanced contractile responses of aortae and mesenteric arteries from diabetic rats to NA was also investigated. The PKC inhibitor, staurosporine abolished the difference in the magnitude of contraction observed between control and diabetic arteries in response to NA. Maximum contractile responses of mesenteric arteries but not aortae or portal veins from diabetic rats to the PKC activator, PDB, were significantly enhanced compared with controls. The enhanced contractile responses of diabetic mesenteric arteries to PDB were abolished in the presence of staurosporine, in the presence of Ca²⁺-channel blockers, or in the absence of extracellular Ca²⁺.
These results indicate that phosphoinositide metabolism is enhanced in aortae and mesenteric arteries from STZ-induced diabetic rats in response to NA, via α₁-adrenoceptor stimulation. The enhancement may contribute to the increased contractile responsiveness of these vessels to NA. In addition, increased activation of PKC-mediated processes, which are dependent on the presence of extracellular Ca²⁺, may further contribute to the enhanced contractile responses of diabetic mesenteric arteries to NA. However, increased influx of extracellular Ca²⁺ by other mechanisms may also contribute to the enhanced contraction of diabetic arteries to submaximum concentrations of NA. Diabetes does not seem to induce such changes in portal veins. === Pharmaceutical Sciences, Faculty of === Graduate
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