Hyperinsulinemia increases sympathetic nerve activity and contributes to cardiovascular dysfunction in obesity and diabetes. Neurons of the hypothalamic paraventricular nucleus regulate sympathetic nerve activity through mono- and poly-synaptic connections to preganglionic neurons in the spinal cord. The purpose of the present study was to determine whether hypothalamic paraventricular nucleus neurons mediate the sympathetic response to insulin. Hyperinsulinemic-euglycemic clamps were performed in α-chloralose-anesthetized, male Sprague-Dawley rats (280–420 g) by an infusion of insulin (3.75 mU/kg/min) and 50% dextrose (0.75–2.0 ml/h) for 120 min. At 90 min, insulin significantly increased lumbar sympathetic nerve activity without any change in renal sympathetic nerve activity, heart rate, or blood glucose levels. Inhibition of the hypothalamic paraventricular nucleus with bilateral injection of the GABAA receptor agonist muscimol completely reversed the sympathoexcitatory response. However, direct injection of insulin into the hypothalamic paraventricular nucleus did not alter lumbar sympathetic nerve activity and thereby suggests insulin activates neurons upstream of the hypothalamic paraventricular nucleus. Interestingly, the sympathetic response to insulin was eliminated by hypothalamic paraventricular nucleus injection of the melancortin 3/4 receptor antagonist SHU9119 but unaffected by the angiotensin II type 1 receptor antagonist losartan. A final set of experiments suggests activation of hypothalamic paraventricular nucleus neurons during hyperinsulinemia increases glutamatergic drive to the rostral ventrolateral medulla. Collectively, these findings indicate insulin activates a melanocortin-dependent pathway to the hypothalamic paraventricular nucleus that increases glutamatergic drive to the rostral ventrolateral medulla and alter cardiovascular function.

Accumulating evidence suggests that insulin acts within the hypothalamus to alter sympathetic nerve activity (SNA) and baroreflex function. Although insulin receptors are widely expressed across the hypothalamus, recent evidence suggests that neurons of the arcuate nucleus (ARC) play an important role in the sympathoexcitatory response to insulin. The purpose of the present study was to determine whether circulating insulin acts directly in the ARC to elevate SNA. In anesthetized male Sprague-Dawley rats (275-425 g), the action of insulin was neutralized by microinjection of an anti-insulin affibody (1 ng/40 nl). To verify the efficacy of the affibody, ARC pretreatment with injection of the anti-insulin affibody completely prevented the increase in lumbar SNA produced by ARC injection of insulin. Next, ARC pretreatment with the anti-insulin affibody attenuated the lumbar sympathoexcitatory response to intracerebroventricular injection of insulin. Third, a hyperinsulinemic-euglycemic clamp increased lumbar, but not renal, SNA in animals that received ARC injection of a control affibody. However, this sympathoexcitatory response was absent in animals pretreated with the anti-insulin affibody in the ARC. Injection of the anti-insulin affibody in the adjacent ventromedial hypothalamus did not alter the sympathoexcitatory response to insulin. The ability of the anti-insulin affibody to prevent the sympathetic effects of insulin cannot be attributed to a general inactivation or nonspecific effect on ARC neurons as the affibody did not alter the sympathoexcitatory response to ARC disinhibition by gabazine. Collectively, these findings suggest that circulating insulin acts within the ARC to increase SNA.

Insulin acts within the central nervous system to regulate food intake and sympathetic nerve activity (SNA). Strong evidence indicates that glucocorticoids impair insulin-mediated glucose uptake and food intake. However, few data are available regarding whether glucocorticoids also modulate the sympathoexcitatory response to insulin. Therefore, the present study first confirmed that chronic administration of glucocorticoids attenuated insulin-induced increases in SNA and then investigated whether these effects were attributed to deficits in central insulin-mediated responses. Male Sprague-Dawley rats were given access to water or a drinking solution of the glucocorticoid agonist dexamethasone (0.3 μg/ml) for 7 days. A hyperinsulinemic-euglycemic clamp significantly increased lumbar SNA in control rats. This response was significantly attenuated in rats given access to dexamethasone for 7, but not 1, days. Similarly, injection of insulin into the lateral ventricle or locally within the arcuate nucleus (ARC) significantly increased lumbar SNA in control rats but this response was absent in rats given access to dexamethasone. The lack of a sympathetic response to insulin cannot be attributed to a generalized depression of sympathetic function or inactivation of ARC neurons as electrical activation of sciatic afferents or ARC injection of gabazine, respectively, produced similar increases in SNA between control and dexamethasone-treated rats. Western blot analysis indicates insulin produced similar activation of Akt Ser(473) and rpS6 Ser(240/244) in the ventral hypothalamus of control and dexamethasone-treated rats. Collectively, these findings suggest that dexamethasone attenuates the sympathoexcitatory actions of insulin through a disruption of ARC neuronal function downstream of Akt or mammalian target of rapamycin (mTOR) signaling.

Insulin contributes to cardiovascular dysfunction by central actions to increase sympathetic nerve activity (SNA). The arcuate nucleus (ARC) abundantly expresses insulin receptors, and direct ARC injection of insulin raises lumbar SNA (LSNA). Therefore, we hypothesized that peripheral insulin acts on ARC neurons to increase LSNA and tested this by using an anti‐insulin affibody to neutralize insulin. To test the efficacy of the affibody, anesthetized Sprague‐Dawley rats received bilateral ARC injection (40nL) of either an anti‐insulin (1μg) or control affibody (1μg) ~10 min before ARC injection of insulin (4μU). The anti‐insulin vs control affibody abolished the increased LSNA (120 min: 99±1% vs 128 ±8%, n=4; P<0.05). Blood pressure and heart rate were similar between groups. Second, ARC injection of anti‐insulin vs control affibody prevented the increased LSNA during a hyperinsulinemic‐euglycemic clamp (7.5mU/kg/min, IV; 114±5% vs 153±15%, n=5–6; P<0.05). Yet, ARC injection of gabazine elevated LSNA in both anti‐insulin vs control animals suggesting the affibody does not generally inhibit ARC neurons. These results indicate circulating insulin acts directly on ARC neurons to increase SNA.Grant Funding Source : HL090826, APS UGSRF