Glutamatergic receptor activation in the rostral ventrolateral medulla mediates the sympathoexcitatory response to hyperinsulinemia

Abstract

Hyperinsulinemia increases sympathetic nerve activity (SNA) and has been linked to cardiovascular morbidity in obesity. The rostral ventrolateral medulla (RVLM) plays a key role in the regulation of SNA and arterial blood pressure (ABP). Many sympathoexcitatory responses are mediated by glutamatergic receptor activation within the RVLM, and both the central renin-angiotensin and melanocortin systems are implicated in the sympathoexcitatory response to hyperinsulinemia. Therefore, we hypothesized that one or more of these neurotransmitters in the RVLM mediate the sympathoexcitatory response to insulin. Hyperinsulinemic-euglycemic clamps were performed in alpha-chloralose anesthetized, male Sprague-Dawley rats by infusion of insulin (3.75 mU/kg per minute, IV) and 50% dextrose solution for 120 minutes. Physiological increases in plasma insulin elevated lumbar SNA, with no change in renal SNA, ABP, or blood glucose. Microinjection of the ionotropic glutamate receptor antagonist kynurenic acid into the RVLM significantly reduced lumbar SNA and ABP. Selective blockade of NMDA but not non-NMDA glutamate receptors resulted in similar reductions of lumbar SNA. In marked contrast, microinjection of the angiotensin II type 1 receptor antagonist losartan or the melanocortin 3/4 antagonist SHU9119 had no effect on lumbar SNA or ABP. Western blot analysis showed that insulin receptor expression is significantly lower in the RVLM than the hypothalamus, and direct microinjection of insulin into the RVLM did not significantly increase lumbar SNA. These findings suggest that hyperinsulinemia increases lumbar SNA by activation of a glutamatergic NMDA-dependent projection to the RVLM.

Figure 1

Plasma insulin concentrations at baseline, 60, and 120 min during a hyperinsulinemic-euglycemic clamp (3.75 mU/kg/min, n=9; 7.5 mU/kg/min, n=3) or saline infusion (n=3). Plasma insulin concentrations from LF (n=5), OR (n=5) and OP (n=6) rats were analyzed for purposes of comparison. Plasma insulin levels were not different between OP rats and control rats infused with 7.5 mU/kg/min. *Significant difference vs baseline levels (P<0.05), †Significant difference versus 3.75 mU/kg/min (P<0.05), ‡Significant difference versus LF and OR rats (P<0.05).

Figure 2

Representative examples of ABP, mean ABP, and lumbar SNA during RVLM microinjection of KYN in rats receiving a (A) hyperinsulinemic-euglycemic clamp or (B) saline infusion. Traces for raw lumbar SNA represent (a) baseline, (b) peak infusion, and (c) post-KYN injection.

Figure 3

Summary data of lumbar SNA, mean ABP and blood glucose during RVLM microinjection of KYN (∇) in rats receiving a hyperinsulinemic-euglycemic clamp (●, n=5) or saline infusion (□, n=5). Injection of KYN significantly reduced lumbar SNA in rats receiving a hyperinsulinemic clamp but had no effect in those receiving saline infusion. *Significant difference vs saline infused-rats (P<0.05), †Significant difference vs pre-injection or 90-min value (P<0.05).

Figure 4

Peak changes in lumbar SNA and mean ABP after bilateral microinjection of aCSF, KYN, AP5, or NBQX into the RVLM during a saline infusion (n=3-7 per group) or hyperinsulemic-euglycemic clamp (n=3-7 per group). *Significant difference vs rats infused with saline within same drug treatment or rats infused with insulin + aCSF (P<0.05)

Figure 5

Peak changes in lumbar SNA and mean ABP after bilateral microinjection of aCSF, KYN, losartan, or SHU 9119 into the RVLM during a saline infusion (n=4-9 per group) or hyperinsulinemic-euglycemic clamp (n=3-6 per group). * Significant difference vs rats infused with saline within same drug treatment or rats infused with insulin + aCSF (P<0.05)

Figure 6

(A) Examples of western blot analysis for insulin receptor β and γ-tubulin in the hypothalamus (H) and RVLM (R). Insulin receptor β expression as a ratio to γ-tubulin was significantly lower in the RVLM versus the hypothalamus (n=4 per group, *P<0.01). (B) Change in lumbar SNA and ABP after injection of insulin into the RVLM (n=3-4 per group) or lateral ventricle (n=5). *Significant difference versus aCSF or 0 insulin (P<0.01).