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Cold-induced vasoconstriction in cutaneous blood vessels is mediated in part by increased activity of vascular smooth muscle α2-adrenoceptors (VSM α2-ARs). In mouse cutaneous arteries, α2C-ARs are normally silent at 37°C but mediate cold-induced augmentation of α2-AR responsiveness. In transfected HEK293 cells, this functional rescue is mediated by cold-induced translocation of α2C-ARs from the Golgi to the plasma membrane. Experiments were performed to determine the role of Rho/Rho kinase signaling in this process. Inhibition of Rho kinase (fasudil, Y27632 or H-1152) did not affect constriction of isolated mouse tail arteries to the α2-AR agonist UK 14 304 at 37°C but dramatically reduced the augmented responses to the agonist at 28°C. After Rho kinase inhibition, cooling no longer increased constriction evoked by α2-AR stimulation. Cooling (to 28°C) activated Rho in VSM cells and increased the calcium sensitivity of constriction in α toxin-permeabilized arteries. Stimulation of α2-ARs in VSM cells had no effect on Rho activity or calcium sensitivity at 37°C or 28°C. In HEK293 cells transfected with α2C-ARs, cooling (to 28°C) stimulated the translocation of α2C-ARs to the plasma membrane and this effect was prevented by inhibition of Rho kinase, using fasudil or RNA interference. Consistent with inhibition of the spatial rescue of α2C-ARs, fasudil inhibited α2-AR–mediated mobilization of calcium in tail arteries at 28°C but not 37°C. Therefore, cold-induced activation of Rho/Rho kinase can mediate cold-induced constriction in cutaneous arteries by enabling translocation of α2C-ARs to the plasma membrane and by increasing the calcium sensitivity of the contractile process.

Cold constricts cutaneous blood vessels by selectively increasing the activity of smooth muscle α₂-adrenoceptors (α₂-ARs). In mouse tail arteries, α₂-AR constriction is mediated by α_2A-ARs at 37°C, whereas the cold-induced augmentation in α₂-AR activity is mediated entirely by α_2C-ARs. Cold causes translocation of α_2C-ARs from the trans-Golgi to the plasma membrane, mediated by cold-induced activation of RhoA and Rho kinase. The present experiments analyzed the mechanisms underlying these responses. Mouse tail arteries were studied in a pressure myograph. Cooling the arteries (28°C) caused a rapid increase in reactive oxygen species (ROS) in smooth muscle cells, determined by confocal microscopy of arteries loaded with the ROS-sensitive probes, dichlorodihydrofluorescein or reduced Mitotracker Red. The inhibitor of mitochondrial complex I rotenone (10 μmol/l), the antioxidant N-acetylcysteine (NAC; 20 mmol/l), or the cell-permeable mimic of superoxide dismutase MnTMPyP (50 μmol/l) did not affect vasoconstriction to α₂-AR stimulation (UK-14304) at 37°C but dramatically inhibited the response at 28°C. Indeed, these ROS inhibitors abolished the cold-induced increase in α₂-AR constrictor activity. NAC (20 mmol/l) or MnTMPyP (50 μmol/l) also abolished the cold-induced activation of RhoA in human cultured vascular smooth muscle cells and the cold-induced mobilization of α_2C-ARs to the cell surface in human embryonic kidney 293 cells transfected with the receptor. The combined results suggest that cold-induced constriction is mediated by redox signaling in smooth muscle cells, initiated by mitochondrial generation of ROS, which stimulate RhoA/Rho kinase signaling and the subsequent mobilization of α_2C-ARs to the cell surface. Altered activity of ROS may contribute to cold-induced vasospasm occurring in Raynaud's phenomenon.

Objective—To determine the metabolic phenotype of a group of laminitis-prone ponies when at pasture in summer, compared with when at pasture in winter. Animals—40 ponies of various breeds predisposed to recurrent pasture-associated laminitis and 40 unaffected control ponies. Procedures—Body condition score and size of the crest of the neck were assessed, blood samples obtained, and blood pressure measured by use of an indirect oscillometric technique, while ponies were kept on winter pasture (last week of November or beginning of December) and again on summer pasture (June). Serum insulin concentration and plasma glucose, triglyceride, uric acid, and ACTH concentrations were measured. Insulin sensitivity was calculated with proxies derived from basal serum insulin and plasma glucose concentrations. Results—No significant differences were apparent between ponies predisposed to laminitis and control ponies during winter. However, in June, laminitis-prone ponies had increased serum insulin concentration and plasma triglyceride and uric acid concentrations, compared with control ponies. Also, laminitis-prone ponies were relatively insulin resistant, compared with control ponies. Mean blood pressure was significantly higher during summer in laminitis-prone ponies (median [interquartile range], 89.6 mm Hg [78.3 to 96.9 mm Hg]), compared with control ponies (76.8 mm Hg [69.4 to 85.2 mm Hg]). Conclusions and Clinical Relevance—Summer pastures appear to induce metabolic responses in some ponies, leading to expression of the prelaminitic phenotype, which includes hypertension as well as insulin resistance. Signs of this metabolic syndrome may not be apparent in affected ponies during periods of grazing winter pasture. Understanding this syndrome may enable improved countermeasures to be devised to prevent laminitis.