Rapid Insulin-Mediated Increase in Microvascular Glycocalyx Accessibility in Skeletal Muscle May Contribute to Insulin-Mediated Glucose Disposal in Rats

Abstract

It has been demonstrated that insulin-mediated recruitment of microvascular blood volume is associated with insulin sensitivity. We hypothesize that insulin rapidly stimulates penetration of red blood cells (RBC) and plasma into the glycocalyx and thereby promotes insulin-mediated glucose uptake by increasing intracapillary blood volume. Experiments were performed in rats; the role of the glycocalyx was assessed by enzymatic degradation using a bolus of hyaluronidase. First, the effect of insulin on glycocalyx accessibility was assessed by measuring the depth of penetration of RBCs into the glycocalyx in microvessels of the gastrocnemius muscle with Sidestream Dark-field imaging. Secondly, peripheral insulin sensitivity was determined using intravenous insulin tolerance tests (IVITT). In addition, in a smaller set of experiments, intravital microscopy of capillary hemodynamics in cremaster muscle and histological analysis of the distribution of fluorescently labeled 40 kDa dextrans (D40) in hindlimb muscle was used to evaluate insulin-mediated increases in capillary blood volume. Insulin increased glycocalyx penetration of RBCs by 0.34±0.44 µm (P<0.05) within 10 minutes, and this effect of insulin was greatly impaired in hyaluronidase treated rats. Further, hyaluronidase treated rats showed a 35±25% reduction in whole-body insulin-mediated glucose disposal compared to control rats. Insulin-mediated increases in capillary blood volume were reflected by a rapid increase in capillary tube hematocrit from 21.1±10.1% to 29.0±9.8% (P<0.05), and an increase in D40 intensity in individual capillaries of 134±138% compared to baseline at the end of the IVITT. These effects of insulin were virtually abolished in hyaluronidase treated animals. In conclusion, insulin rapidly increases glycocalyx accessibility for circulating blood in muscle, and this is associated with an increased blood volume in individual capillaries. Hyaluronidase treatment of the glycocalyx abolishes the effects of insulin on capillary blood volume and impairs insulin-mediated glucose disposal.

Acute enzymatic glycocalyx degradation results in reduced insulin sensitivity but normal glucose tolerance in conscious rats

“…In a previous study [6], we showed in anesthetized rats that acute enzymatic glycocalyx degradation is associated with a decrease in insulin-mediated glucose disposal, suggesting a role for the glycocalyx in regulation of insulin sensitivity. In the current study, we similarly demonstrated this role in conscious rats and showed that the evoked insulin resistance did not affect glucose tolerance because of increased pancreatic insulin release in response to an exogenous glucose load.…”

“…Insulin tolerance tests in our previous study were performed without the use of somatostatin and during isoflurane anesthesia [6], and it has been shown previously that isoflurane anesthesia affects glucose utilization [17,18]. Therefore, in the current study, we measured glucose and insulin tolerance in conscious rats 1 week after cannulation.…”

“…The endothelial glycocalyx, which shields the vascular wall from direct exposure to the flowing blood, has, since the last decade, been indicated to play an important role in the protection of the endothelium against atherogenic insults [4,5], whereas we recently provided evidence suggesting that this compartment may also be involved in the regulation of insulin sensitivity. In anesthetized rats, we showed that insulin, by diminishing the barrier properties of the glycocalyx and increasing its accessibility for the flowing blood, rapidly increased microvascular blood volume in muscles [6], an action that has been indicated in studies by Rattigan and colleagues [7][8][9] to contribute to insulin-mediated glucose disposal. Indeed, we observed that acute enzymatic degradation of the glycocalyx in the rats by intravenous administration of hyaluronidase was associated with an acute decrease in whole-body insulin-mediated glucose disposal during an intravenous insulin tolerance test [6].…”

“…In anesthetized rats, we showed that insulin, by diminishing the barrier properties of the glycocalyx and increasing its accessibility for the flowing blood, rapidly increased microvascular blood volume in muscles [6], an action that has been indicated in studies by Rattigan and colleagues [7][8][9] to contribute to insulin-mediated glucose disposal. Indeed, we observed that acute enzymatic degradation of the glycocalyx in the rats by intravenous administration of hyaluronidase was associated with an acute decrease in whole-body insulin-mediated glucose disposal during an intravenous insulin tolerance test [6]. The physiological relevance of these latter observations for glucose regulation may, however, be questioned, as the insulin tolerance test was performed during isoflurane anesthesia, requiring a supraphysiological dose of insulin to be administered.…”

“…Although traditionally the insulinmediated increase in capillary blood volume was explained by an increase in the number of perfused capillaries (capillary recruitment) [7], we recently showed in anesthetized rats that insulin can also increase blood volume within already perfused microvessels by increasing blood accessibility into the glycocalyx, and that this vascular action of insulin is impaired after enzymatic glycocalyx degradation. Further, it was suggested that this impaired insulin-mediated microvascular blood volume increase in muscles underlay the B30% reduction in the glucose disposal rate in the IVITT after hyaluronidase treatment in the rats [6]. To check whether the reduction in glucose disposal from the blood was due to diminished insulin action in skeletal muscle, Akt phosphorylation (expressed as the Glycocalyx contributes to insulin sensitivity Eskens et al 71 ratio of phosphorylated Akt to total Akt) was measured in the soleus muscle after the IVITT in a few of these anesthetized rats (n = 5, unpaired measurements).…”