Acute elevation of circulating fatty acids impairs downstream insulin signalling in rat skeletal muscle in vivo independent of effects on stress signalling

Abstract

The aim of this study was to examine the effect of an acute, physiological increase in plasma free fatty acid (FFA) on initial signalling events in rat red quadriceps muscle (RQ). Male Wistar rats received a 7% glycerol (GLYC) or 7% Intralipid/heparin (LIP) infusion for 3 h, after which they were either killed or infused with insulin at a rate of 0 . 5 U/kg per h for 5 min, before RQ collection. Plasma FFAs were elevated to w2 mM in the LIP rats only. Insulin-stimulated insulin receptor (IR) Tyr1162/Tyr1163 phosphorylation and IR substrate (IRS)-1 Tyr612 phosphorylation were increased at least twofold over basal in GLYC rats with insulin and this increase was not significantly impaired in the LIP rats. However, there was no insulin-stimulated protein kinase B (PKB) Ser473 or glycogen synthase kinase (GSK)-3b Ser9 phosphorylation in the LIP rats, compared with at least a twofold increase over basal in GLYC rats for both proteins. c-Jun N-terminal kinase, inhibitor of k kinase b and inhibitor of nuclear factor-kB phosphorylation and total protein expression, as well as Ser307-IRS-1 phosphorylation, were not altered by lipid infusion compared with GLYC infusion. These data indicate that acute, physiological elevation in FFA has a greater impact on insulin signalling downstream of IR and IRS-1, at the level of PKB and GSK-3b, and that under these conditions stress signalling pathways are not significantly stimulated. Decreased PKB and GSK-3b phosphorylation in RQ may therefore be primary determinants of the reduced insulin action observed in situations of acute FFA oversupply.

Lipid and insulin infusion-induced skeletal muscle insulin resistance is likely due to metabolic feedback and not changes in IRS-1, Akt, or AS160 phosphorylation

“…Furthermore, defects in activation of components of the insulinsignaling pathway have been linked with skeletal muscle in- sulin resistance. These defects include reduced tyrosine phosphorylation of IRS-1, which is thought to be due to increased inhibitory serine phosphorylation, reduced phosphatidylinositol 3-kinase activity, reduced Akt1 (but not Akt2 or Akt3) activation, decreased PKC/ activation, and lower glycogen synthase activity and phosphorylation (11,17,29,30,50). Accumulation of active lipid intermediates, such as DAG, ceramide, and LCACoA, has been linked with insulin resistance (5,8,20,53,56).…”

“…Degradation of IRS-1 protein levels has also been suggested as a process whereby insulin signaling is reduced in insulin-resistant states (57). Inhibition of Akt activation is thought to be due to increased activation of the protein phosphatase 2A (45) or PKC/ (40) by accumulated ceramide.The majority of in vivo studies that have implicated defects in insulin signaling as a mechanism for insulin resistance have done so in models of hyperlipidemia, including high-fat feeding (12, 55, 58) and acute lipid infusion (11,18,20,50). Additional work has shown that incubation of isolated muscles or cultured myotubes with high fatty acid [predominantly palmitate (C 16 )] concentrations leads to an impairment in insulin signaling and reduced insulin action (1,19,56).…”

“…The majority of in vivo studies that have implicated defects in insulin signaling as a mechanism for insulin resistance have done so in models of hyperlipidemia, including high-fat feeding (12,55,58) and acute lipid infusion (11,18,20,50). Additional work has shown that incubation of isolated muscles or cultured myotubes with high fatty acid [predominantly palmitate (C 16 )] concentrations leads to an impairment in insulin signaling and reduced insulin action (1,19,56).…”

“…Furthermore, a recent in vitro study reported that, in isolated soleus muscle, 6 h of palmitate incubation reduced acute insulin-stimulated glucose uptake and membrane GLUT4 protein, did not change Akt phosphorylation (Thr 308 and Ser 473 ), and reduced AS160 phosphorylation (1). In the majority of investigations of the mechanisms associated with lipid oversupply-induced insulin resistance in skeletal muscle, insulin was not elevated from the commencement of the lipid infusion (3,8,11,16,18,20,29,30,49,50,52). This is important, inasmuch as hyperlipidemia is often associated with hyperinsulinemia in human subjects with type 2 diabetes (4).…”

Fatty acid metabolism, energy expenditure and insulin resistance in muscle

“…While there are many studies showing clear differences in the phosphorylation status of various insulin signalling proteins after insulin stimulation in control and FAexposed or obese or high-fat diet-fed muscle, these changes are not always consistent. For example, a change in Akt phosphorylation is not always accompanied by a detectable change in downstream GSK3 or AS160 phosphorylation or upstream changes in IRS1 phosphorylation (Frangioudakis & Cooney 2008, Hoehn et al 2008, Tonks et al 2013. There are a number of studies reporting that insulin-stimulated Akt activation is in fact not impaired in the muscle of obese individuals with insulin resistance, of glucose-intolerant first-degree relatives of patients with T2D and of patients with T2D (Kim et al 1999, Storgaard et al 2004.…”