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OBJECTIVE--Glucokinase (GCK) acts as a component of the 'glucose sensor' in pancreatic [beta]-cells and possibly in other tissues, including the brain. However, >99% of GCK in the body is located [...]

We previously showed that insulin has a profound effect to suppress pyruvate dehydrogenase kinase (PDK) 4 expression in rat skeletal muscle. In the present study, we examined whether insulin's effect on PDK4 expression is impaired in acute insulin-resistant states and, if so, whether this change is accompanied by decreased insulin's effects to stimulate Akt and forkhead box class O (FOXO) 1 phosphorylation. To induce insulin resistance, conscious overnight-fasted rats received a constant infusion of Intralipid or lactate for 5 h, while a control group received saline infusion. Following the initial infusions, each group received saline or insulin infusion (n = 6 or 7 each) for an additional 5 h, while saline, Intralipid, or lactate infusion was continued. Plasma glucose was clamped at basal levels during the insulin infusion. Compared with the control group, Intralipid and lactate infusions decreased glucose infusion rates required to clamp plasma glucose by ~60% (P < 0.01), confirming the induction of insulin resistance. Insulin's ability to suppress PDK4 mRNA level was impaired in skeletal muscle with Intralipid and lactate infusions, resulting in two- to threefold higher PDK4 mRNA levels with insulin (P < 0.05). Insulin stimulation of Akt and FOXO1 phosphorylation was also significantly decreased with Intralipid and lactate infusions. These data suggest that insulin's effect to suppress PDK4 gene expression in skeletal muscle is impaired in insulin-resistant states, and this may be due to impaired insulin signaling for stimulation of Akt and FOXO1 phosphorylation. Impaired insulin's effect to suppress PDK4 expression may explain the association between PDK4 overexpression and insulin resistance in skeletal muscle., Pyruvate dehydrogenase kinase (PDK) phosphorylates and inactivates pyruvate dehydrogenase complex that catalyzes a rate-limiting step of glucose oxidation, i.e., the conversion of pyruvate to acetyl CoA. Previous studies (1-3) showed [...]

Insulin stimulates both glucose and [K.sup.+] uptake, and high-fat feeding is known to decrease insulin-stimulated glucose uptake. The purpose of this study was to examine whether insulin's actions on glucose [...]

To examine whether impairment of intracellular glucose metabolism precedes insulin resistance, we determined the time courses of changes in insulin-stimulated glucose uptake, glycolysis, and glycogen synthesis during high-fat feeding in [...]

The effects of elevated plasma free fatty acid (FFA) levels on insulin-stimulated whole-body and skeletal muscle glucose transport, glucose uptake, glycolysis, and glycogen synthesis were studied in conscious rats during [...]

To determine the relative time courses of changes in peripheral and hepatic insulin action and skeletal muscle GLUT4 protein levels after a streptozotocin (STZ) injection in rats, we performed hyperinsulinemic [...]

Effects of 24-h and 48-h fasting on maximal insulin-stimulated whole-body and muscle glucose uptake, glycogen synthesis, and glycolysis were studied in conscious rats by combining the glucose clamp technique with [...]

Pancreatic β-Cell Hyperfunction With Normal Insulin Sensitivity We used intravenous glucose tolerance tests in vivo and 3-O-methylglucose transport into skeletal muscle in vitro to assess glucose tolerance, pancreatic β-cell function, [...]

To evaluate the role of the hexosamine biosynthesis pathway (HBP) in fat-induced insulin resistance, we examined whether fat-induced insulin resistance is additive to that induced by increased HBP flux via [...]

The hexosamine biosynthesis pathway (HBP) has been implicated in hyperglycemia-induced insulin resistance. The HBP may also be involved in the development of insulin resistance at euglycemia with suppression of glucose [...]

The mechanism of insulin's suppression of endogenous glucose production (EGP) is not well understood. In vivo studies suggest that inhibition of either gluconeogenesis or glycogenolysis alone does not result in [...]

Previous studies showed that lactate infusion induces peripheral insulin resistance, but the underlying mechanisms remain unclear. There is substantial evidence that impairment of intracellular glucose metabolism precedes and causes peripheral [...]

We examined the individual and combined effects of high-fat diet (HFD) and exercise training on insulin-stimulated whole body glucose fluxes and several key steps of glucose metabolism in skeletal muscle. [...]

Dietary potassium loading results in rapid kaliuresis, natriuresis, and diuresis associated with reduced phosphorylation (p) of the distal tubule Na+-Cl−cotransporter (NCC). Decreased NCC-p inhibits NCC-mediated Na+reabsorption and shifts Na+downstream for reabsorption by epithelial Na+channels (ENaC), which can drive K+secretion. Whether the signal is initiated by ingesting potassium or a rise in plasma K+concentration ([K+]) is not understood. We tested the hypothesis, in male rats, that an increase in plasma [K+] is sufficient to reduce NCC-p and drive kaliuresis. After an overnight fast, a single 3-h 2% potassium (2%K) containing meal increased plasma [K+] from 4.0 ± 0.1 to 5.2 ± 0.2 mM; increased urinary K+, Na+, and volume excretion; decreased NCC-p by 60%; and marginally reduced cortical Na+-K+-2Cl−cotransporter (NKCC) phosphorylation 25% (P= 0.055). When plasma [K+] was increased by tail vein infusion of KCl to 5.5 ± 0.1 mM over 3 h, significant kaliuresis and natriuresis ensued, NCC-p decreased by 60%, and STE20/SPS1-related proline alanine-rich kinase (SPAK) phosphorylation was marginally reduced 35% (P= 0.052). The following were unchanged at 3 h by either the potassium-rich meal or KCl infusion: Na+/H+exchanger 3 (NHE3), NHE3-p, NKCC, ENaC subunits, and renal outer medullary K+channel. In summary, raising plasma [K+] by intravenous infusion to a level equivalent to that observed after a single potassium-rich meal triggers renal kaliuretic and natriuretic responses, independent of K+ingestion, likely driven by decreased NCC-p and activity sufficient to shift sodium reabsorption downstream to where Na+reabsorption and flow drive K+secretion.

Extracellular K+homeostasis has been explained by feedback mechanisms in which changes in extracellular K+concentration drive renal K+excretion directly or indirectly via stimulating aldosterone secretion. However, this cannot explain meal-induced kaliuresis, which often occurs without increases in plasma K+or aldosterone concentrations. Recent studies have produced evidence supporting a feedforward control in which gut sensing of dietary K+increases renal K+excretion (and extrarenal K+uptake) independent of plasma K+concentrations, namely, a gut factor. This review focuses on these new findings and discusses the role of gut factor in acute and chronic regulation of extracellular K+as well as in the beneficial effects of high K+intake on the cardiovascular system.

We tested the hypothesis that K+intake is sensed by putative K+sensors in the splanchnic areas, and renal K+handling is regulated by this signal. K+was infused for 2 h into overnight-fasted rats via the jugular vein (systemic infusion), hepatic portal vein (intraportal infusion), or stomach (intragastric infusion) (n= 5 each), and plasma K+concentration ([K+]) and renal K+excretion were measured during the 2-h preinfusion, 2-h K+infusion, and 3-h washout periods. During systemic K+infusion, plasma [K+] increased by ∼1.3 mM (P< 0.05), and, on cessation of the K+infusion, plasma [K+] fell to the preinfusion level within 1–2 h. Renal K+excretion changed in proportion to the changes in plasma [K+]. During intraportal or intragastric K+infusion, plasma [K+] and renal K+excretion profiles were similar to those with systemic infusion. The effects of K+infusions via the different routes (n= 5 or 6 each) were also studied during simultaneous feeding of overnight-fasted rats with a K+-deficient diet. During the meal, intraportal infusion resulted in increases in plasma [K+] similar to those with the systemic K+infusion, while intragastric K+infusion did not significantly increase plasma [K+]. Thus, when the intragastric K+infusion was combined with a meal, there was marked enhancement of clearance of the K+infused, which was associated with an apparent increase in renal efficiency of K+excretion. These data suggest that there may be a gut factor that enhances renal efficiency of K+excretion during meal (or dietary K+) intake.

There is a positive association between diets rich in potassium, control of blood pressure, and prevention of stroke. Extracellular [K+] is regulated closely to maintain normal membrane excitability by the concerted regulatory responses of muscle and kidney. Although kidney is responsible for ultimately matching K+output to K+intake each day, muscle contains more than 90% of the body’s K+and can buffer changes in extracellular fluid [K+] by either acutely taking up extracellular fluid K+or releasing intracellular fluid K+from muscle. It long has been assumed that the changes in muscle K+transport are a function of sodium pump (Na,K-adenosine triphosphatase [Na, K-ATPasel]) abundance, especially that of the a2 isoform, which predominates in skeletal muscle. To test the physiologic significance of changes in muscle Na,K-ATPase expression, we developed the K+clamp, which measures insulin-stimulated cellular K+uptake in vivo in the conscious rat. By using the K+clamp we discovered that significant insulin resistance to cell K+uptake occurs as follows: (1) early in K+deprivation before a decrease in muscle sodium pump pool size, and (2) during glucocorticoid treatment, which increases muscle Na,K-ATPase a2 levels greater than 50%. We also discovered that adaptation of renal and extrarenal K+handling to altered K+balance often occurs without changes in plasma [K+], supporting a feedforward mechanism involving K+sensing in the splanchnic bed and adjustment of K+handling. These findings establish the advantage of combining molecular analyses of Na,K-ATPase expression and activity with systems analyses of cellular K+uptake and excretion in vivo to reveal regulatory mechanisms operating to control K+homeostasis.

Maintaining extracellular fluid (ECF) K+concentration ([K+]) within a narrow range is accomplished by the concerted responses of the kidney, which matches K+excretion to K+intake, and skeletal muscle, the main intracellular fluid (ICF) store of K+, which can rapidly buffer ECF [K+]. In both systems, homologous P-type ATPase isoforms are key effectors of this homeostasis. During dietary K+deprivation, these P-type ATPases are regulated in opposite directions: increased abundance of the H,K-ATPase “colonic” isoform in the renal collecting duct drives active K+conservation while decreased abundance of the plasma membrane Na,K-ATPase α2-isoform leads to the specific shift of K+from muscle ICF to ECF. The skeletal muscle response is isoform and muscle specific: α2and β2, not α1and β1, levels are depressed, and fast glycolytic muscles lose >90% α2, whereas slow oxidative muscles lose ∼50%; however, both muscle types have the same fall in cellular [K+]. To understand the physiological impact, we developed the “K+clamp” to assess insulin-stimulated cellular K+uptake in vivo in the conscious rat by measuring the exogenous K+infusion rate needed to maintain constant plasma [K+] during insulin infusion. Using the K+clamp, we established that K+deprivation leads to near-complete insulin resistance of cellular K+uptake and that this insulin resistance can occur before any decrease in plasma [K+] or muscle Na+pump expression. These studies establish the advantage of combining molecular analyses of P-type ATPase expression with in vivo analyses of cellular K+uptake and excretion to determine mechanisms in models of disrupted K+homeostasis.

We aimed to test the feasibility of quantifying insulin action on cellular K+uptake in vivo in the conscious rat by measuring the exogenous K+infusion rate needed to maintain constant plasma K+concentration ([K+]) during insulin infusion. In this “K+clamp” the K+infusion rate required to clamp plasma [K+] is a measure of insulin action to increase net plasma K+disappearance. K+infusion rate required to clamp plasma [K+] was insulin dose dependent. Renal K+excretion was not significantly affected by insulin at a physiological concentration (∼90 μU/ml,P> 0.05), indicating that most of insulin-mediated plasma K+disappearance was due to K+uptake by extrarenal tissues. In rats deprived of K+for 2 days, plasma [K+] fell from 4.2 to 3.8 mM, insulin-mediated plasma glucose clearance was normal, but insulin-mediated plasma K+disappearance decreased to 20% of control, even though there was no change in muscle Na-K-ATPase activity or expression, which is believed to be the main K+uptake route. After 10 days K+deprivation, plasma [K+] fell to 2.9 mM, insulin-mediated K+disappearance decreased to 6% of control (glucose clearance normal), and there were 50% decreases in Na-K-ATPase activity and α2-subunit levels. In conclusion, the present study proves the feasibility of the K+clamp technique and demonstrates that short-term K+deprivation leads to a near complete insulin resistance of cellular K+uptake that precedes changes in muscle sodium pump expression.

Recent studies have suggested that the insulin receptor tyrosine kinase inhibitor, membrane glycoprotein PC-1, may play a role in certain insulin resistant states. In the present study, we examined whether either insulin receptor function or PC-1 activity was altered during the development of insulin resistance that occurs with high fat feeding in normal rats. Over the course of 14 days of high fat feeding, both maximal and submaximal (physiological) insulin-stimulated skeletal muscle glucose uptake decreased gradually; after 14 days of high fat feeding, submaximal and maximal insulin-stimulated glucose uptake decreased by ∼40 and ∼50%, respectively. In contrast, in the same muscles (tibialis anterior) of these animals, neither insulin receptor content nor insulin-stimulated insulin receptor autophosphorylation was altered after 14 days of high fat feeding. PC-1 has both nucleotide pyrophosphatase (EC 3.6.1.9) and alkaline phosphodiesterase I (EC 3.1.4.1) enzyme activities. These enzyme activities showed no changes during the course of 14 days of high fat feeding. Individual data revealed that there was no significant correlation between insulin-stimulated glucose uptake and alkaline phosphodiesterase or nucleotide pyrophosphatase activity (P> 0.05). Together, these data indicate that neither defects in insulin receptor function nor elevated PC-1 activities are involved in the development of insulin resistance in rats with high fat feeding, and the insulin resistance induced with high fat feeding is likely due to postreceptor defects in skeletal muscle.