Richard J. Koletsky

Decreased transport of leptin across the blood–brain barrier in rats lacking the short form of the leptin receptor☆ ☆

Leptin is produced in adipose tissue in the periphery, but its satiety effect is exerted in the CNS that it reaches by a saturable transport system across the blood-brain barrier (BBB). The short form of the leptin receptor has been hypothesized to be the transporter, with impaired transport of leptin being implicated in obesity. In Koletsky rats, the splice variant that gives rise to the short form of the leptin receptor contains a point mutation that results in marked obesity. We studied the transport of leptin across the BBB in Koletsky rats and found it to be significantly less than in their lean littermates. By contrast, Sprague-Dawley rats matched in weight to each of these two groups showed no difference in the blood-to-brain influx of leptin. HPLC showed that most of the leptin crossing the BBB in rats remained intact and capillary depletion showed that most of the leptin reached the parenchyma of the brain. The results indicate that the short form of the leptin receptor is involved in the transport of leptin across the BBB.

The I1-imidazoline receptor: from binding site to therapeutic target in cardiovascular disease

Objective To review previous work and present additional evidence characterizing the I1-imidazoline receptor and its role in cellular signaling, central cardiovascular control, and the treatment of metabolic syndromes. Second-generation centrally-acting antihypertensives inhibit sympathetic activity mainly via imidazoline receptors, whereas first-generation agents act via α2-adrenergic receptors. The I1subtype of imidazoline receptor resides in the plasma membrane and binds central antihypertensives with high affinity. Methods and results Radioligand binding assays have characterized I1-imidazoline sites in the brainstem site of action for these agents in the rostral ventrolateral medulla. Binding affinity at I1-imidazoline sites, but not at other classes of imidazoline binding sites, correlates closely with the potency of central antihypertensive agents in animals and in human clinical trials. The antihypertensive action of systemic moxonidine is eliminated by the I1α2-antagonist efaroxan, but not by selective blockade of α2-adrenergic receptors. Until now, the cell signaling pathway coupled to I1-imidazoline receptors was unknown. Using a model system lacking α2-adrenergic receptors (PC12 pheochromocytoma cells) we have found that moxonidine acts as an agonist at the cell level and I1-imidazoline receptor activation leads to the production of the second messenger diacylglycerol, most likely through direct activation of phosphatidylcholine-selective phospholipase C. The obese spontaneously hypertensive rat (SHR; SHROB strain) shows many of the abnormalities that cluster in human syndrome X, including elevations in blood pressure, serum lipids and insulin. SHROB and their lean SHR littermates were treated with moxonidine at 8 mg/kg per day. SHROB and SHR treated with moxonidine showed not only lowered blood pressure but also improved glucose tolerance and facilitated insulin secretion in response to a glucose load. Because α2-adrenergic agonists impair glucose tolerance, I1-imidazoline receptors may contribute to the multiple beneficial effects of moxonidine treatment. Conclusion The I1-imidazoline receptor is a specific high- affinity binding site corresponding to a functional cell-sur-face receptor mediating the antihypertensive actions of moxonidine and other second-generation centrally-acting agents, and may play a role in countering insulin resistance in an animal model of metabolic syndrome X.

Reduced insulin receptor signaling in the obese spontaneously hypertensive Koletsky rat

Insulin resistance is associated with both obesity and hypertension. However, the cellular mechanisms of insulin resistance in genetic models of obese-hypertension have not been identified. The objective of the present study was to investigate the effects of genetic obesity on a background of inherited hypertension on initial components of the insulin signal transduction pathway and glucose transport in skeletal muscle and liver. Oral glucose tolerance testing in SHROB demonstrated a sustained postchallenge elevation in plasma glucose at 180 and 240 min compared with lean spontaneously hypertensive rat (SHR) littermates, which is suggestive of glucose intolerance. Fasting plasma insulin levels were elevated 18-fold in SHROB. The rate of insulin-stimulated 3- O-methylglucose transport was reduced 68% in isolated epitrochlearis muscles from the SHROB compared with SHR. Insulin-stimulated tyrosine phosphorylation of the insulin receptor β-subunit and insulin receptor substrate-1 (IRS-1) in intact skeletal muscle of SHROB was reduced by 36 and 23%, respectively, compared with SHR, due primarily to 32 and 60% decreases in insulin receptor and IRS-1 protein expression, respectively. The amounts of p85α regulatory subunit of phosphatidylinositol-3-kinase and GLUT-4 protein were reduced by 28 and 25% in SHROB muscle compared with SHR. In the liver of SHROB, the effect of insulin on tyrosine phosphorylation of IRS-1 was not changed, but insulin receptor phosphorylation was decreased by 41%, compared with SHR, due to a 30% reduction in insulin receptor levels. Our observations suggest that the leptin receptor mutation fak imposed on a hypertensive background results in extreme hyperinsulinemia, glucose intolerance, and decreased expression of postreceptor insulin signaling proteins in skeletal muscle. Despite these changes, hypertension is not exacerbated in SHROB compared with SHR, suggesting these metabolic abnormalities may not contribute to hypertension in this model of Syndrome X.Insulin resistance is associated with both obesity and hypertension. However, the cellular mechanisms of insulin resistance in genetic models of obese-hypertension have not been identified. The objective of the present study was to investigate the effects of genetic obesity on a background of inherited hypertension on initial components of the insulin signal transduction pathway and glucose transport in skeletal muscle and liver. Oral glucose tolerance testing in SHROB demonstrated a sustained postchallenge elevation in plasma glucose at 180 and 240 min compared with lean spontaneously hypertensive rat (SHR) littermates, which is suggestive of glucose intolerance. Fasting plasma insulin levels were elevated 18-fold in SHROB. The rate of insulin-stimulated 3-O-methylglucose transport was reduced 68% in isolated epitrochlearis muscles from the SHROB compared with SHR. Insulin-stimulated tyrosine phosphorylation of the insulin receptor beta-subunit and insulin receptor substrate-1 (IRS-1) in intact skeletal muscle of SHROB was reduced by 36 and 23%, respectively, compared with SHR, due primarily to 32 and 60% decreases in insulin receptor and IRS-1 protein expression, respectively. The amounts of p85 alpha regulatory subunit of phosphatidylinositol-3-kinase and GLUT-4 protein were reduced by 28 and 25% in SHROB muscle compared with SHR. In the liver of SHROB, the effect of insulin on tyrosine phosphorylation of IRS-1 was not changed, but insulin receptor phosphorylation was decreased by 41%, compared with SHR, due to a 30% reduction in insulin receptor levels. Our observations suggest that the leptin receptor mutation fak imposed on a hypertensive background results in extreme hyperinsulinemia, glucose intolerance, and decreased expression of postreceptor insulin signaling proteins in skeletal muscle. Despite these changes, hypertension is not exacerbated in SHROB compared with SHR, suggesting these metabolic abnormalities may not contribute to hypertension in this model of Syndrome X.

Molecular pathology in the obese spontaneous hypertensive Koletsky rat: a model of syndrome X.

ABSTRACT: The SHROB rat is a unique strain with genetic obesity, hypertriglyceridemia, hyperinsulinemia, renal disease with proteinuria, and genetically determined hypertension, characteristics paralleling human Syndrome X. The obese phenotype results from a single homozygous recessive trait, designated faK, and is allelic with the Zucker fatty trait (fa), but of distinct origin. The faK mutation is a premature stop codon in the extracellular domain of the leptin receptor, resulting in a natural receptor knockout. The SHROB are glucose intolerant compared to heterozygous or wild‐type SHR, but retain fasting euglycemia even on a high sucrose diet, suggesting that diabetes requires polygenic interaction with additional modifier genes. Insulin‐stimulated phosphorylation of tyrosine residues on the insulin receptor and on the associated docking protein IRS‐1 are reduced in skeletal muscle and liver compared to SHR, due mainly to diminished expression of insulin receptor and IRS‐1 proteins. Despite multiple metabolic derangements and severe insulin resistance, hypertension is not exacerbated in SHROB compared to SHR. Thus, insulin resistance and hypertension are independent in this model. Increased activity of the sympathetic nervous system may be a common factor leading by separate pathways to hypertension and to insulin resistance. We studied the chronic effects of sympathetic inhibition with moxonidine on glucose metabolism in SHROB. Moxoni‐dine (8 mg/kg/day), a selective I1‐imidazoline receptor agonist, not only reduced blood pressure but also ameliorated glucose intolerance. Moxonidine reduced fasting insulin by 47% and plasma free fatty acids by 30%. Moxonidine enhanced expression and insulin‐stimulated phosphorylation of IRS‐1 in skeletal muscle by 74 and 27%, respectively. Thus, central sympatholytic therapy not only counters hypertension but also insulin resistance, glucose tolerance, and hyperlipidemia in the SHROB model of Syndrome X.

Sympathetic Nervous System in Salt-Sensitive and Obese Hypertension: Amelioration of Multiple Abnormalities by a Central Sympatholytic Agent

SummaryExcess activity of the sympathetic nervous system (SNS) is linked to human obese hypertension and to salt-sensitive hypertension. Paradoxically, reduced SNS activity has been implicated as a contributor to obesity, particularly in animal models, and salt loading usually inhibits SNS activity. We have investigated the relationship between SNS activity, diet, and hypertension in the obese spontaneously hypertensive rat (SHROB), a model with a recessive obesity trait superimposed on a hypertensive background with multiple metabolic abnormalities resembling human syndrome X. We examined the role of SNS overactivity in the adverse impact of excess dietary salt and the possible beneficial effects of sympatholytic therapy. Mean blood pressure (MBP) was increased in SHROB and SHR fed a 4% NaCl diet. The pressor effect of dietary salt was abolished by ganglionic blockade, suggesting that increased SNS activity contributed to the pressor effect of the high-salt diet. Moxonidine, a second-generation central antihypertensive, controlled hypertension in both SHROB and SHR. Kidney damage in SHROB was accelerated by dietary salt and was reduced by moxonidine. Moxonidine elicited progressive weight loss in SHROB but not in SHR. Food intake in SHROB was reduced to the level of lean SHR. SHROB and SHR treated with moxonidine showed improved glucose tolerance. Additionally, SHROB showed reduced levels of triglycerides, cholesterol, and insulin following moxonidine therapy. Inhibition of the SNS, as with moxonidine therapy, may ameliorate multiple abnormalities and have therapeutic advantages in obese hypertensive syndromes.

Biomedical Rationale for a Wellness Approach to Obesity: An Alternative to a focus on Weight Loss

The direct medical hazards of obesity, although real, have been overstated. Because current remedies for obesity have little long-term effectiveness, no controlled clinical trial has demonstrated improved longevity after weight loss. In contrast, advances in drug therapy for diabetes, hypertension, and high cholesterol allow obese persons affected by these conditions to live healthier lives. Furthermore, weight cycling may cause much of the cardiovascular risk associated with obesity. Repeated loss and regain of weight increases human deaths from heart disease, and in obese laboratory animals weight cycling increases blood pressure, enlarges the heart, damages the kidney, increases abdominal fat deposits, and promotes further weight gain. Additional health risks in obesity may be caused by hazardous treatments for obesity, as illustrated by heart disease caused by diet pills. Obese patients often lack full access to medical services owing in part to social stigma and low self-esteem, which impair self-care activities, and the bias of health professionals. These barriers, along with the prevalence of poverty among the obese, may contribute to the association of obesity with poor health. Medical beliefs about obesity are shaped by expert panels that are highly selective in the data they consider. Experts included on government consensus panels have been disproportionately drawn from the ranks of diet clinic directors, which might explain the congruence between panel recommendations and the economic interests of the diet industry. One remedy is a wellness approach focused on healthy lifestyle, positive attitude to health and self-care, and a disregarding of predetermined weight standards in favor of preventing further weight gain and reducing risk factors. Medical conditions common in obese patients, including hypertension, type-2 diabetes, hyperlipidemia, and sleep apnea, are dealt with directly and aggressively rather than relying on weight loss as the primary treatment. This new approach should improve the physical and mental well being of obese patients.

Refeeding hypertension in obese spontaneously hypertensive rats.

Very-low-calorie diets lower blood pressure acutely in obese humans and rats. However, refeeding after dietary restriction produces mild hypertension in rats. Refeeding hypertension was characterized in genetically obese spontaneously hypertensive rats (obese SHR, Koletsky rat), a model of genetic obesity and hypertension. Obese SHR were fed a restricted diet (Optifast) for 12 days, refed ad libitum for 28 days, dieted again for 12 days, and then refed 4 days and killed. Control obese SHR and lean SHR littermates were fed ad libitum continuously. Dietary restriction led to rapid weight loss followed by prompt regain to baseline weight after return to unrestricted food intake. Heart rate fell with institution of the low-calorie diet and returned to baseline on refeeding. Blood pressure became elevated during refeeding in dieted obese SHR relative to ad libitum fed obese SHR controls. The fall in blood pressure after ganglionic blockade with chlorisondamine was exaggerated in refed obese SHR, and cardiac beta-adrenergic receptors were downregulated. Both of these findings imply increased sympathetic tone. The left ventricular wall was thicker in the refed obese SHR than in the ad libitum fed obese SHR. Shorter cycles of weight loss and regain in lean SHR led to transient increases in blood pressure and heart rate. Cycles of dietary restriction and refeeding in obese SHR elicit sustained blood pressure elevation via sympathetic activation and exacerbate cardiac hypertrophy. Drastic fluctuations in nutrient intake may not be advantageous in hypertension.

Anti-hyperglycemic activity of moxonidine: metabolic and molecular effects in obese spontaneously hypertensive rats.

Hypertension and insulin resistance are often part of a complex set of abnormalities including obesity, hyperlipidemia, and glucose intolerance, described as syndrome X. Besides a common genetic basis, insulin resistance and hypertension might be linked by excessive activity of the sympathetic nervous system. We studied the effects of chronic inhibition of sympathetic activity with the antihypertensive agent moxonidine on glucose metabolism in the genetically obese SHR Koletsky rat (SHROB), a unique animal model which closely resembles human syndrome X, expressing genetic obesity, hypertension, and hyperlipidemia. Moxonidine, a selective I1-imidazoline receptor agonist, was administered to SHROB and SHR for 90 days in food at 8 mg/kg/day. Moxonidine not only lowered blood pressure, but also reduced fasting insulin levels by 49% in SHROB, and reduced plasma free fatty acids by 30%. In lean SHR, moxonidine treatment decreased circulating free fatty acids by 33% compared to controls. During oral glucose tolerance tests, blood glucose levels in moxonidine-treated SHROB were reduced from 60 min onwards, and there was a sharply higher insulin secretion post-challenge compared to control SHROB. Western blot analysis of insulin signaling proteins showed that IRS-1 was decreased 42% in control SHROB compared with SHR. Moxonidine treatment enhanced the expression of IRS-1 protein in skeletal muscle by 74% in SHROB and 40% in SHR. Moxonidine increased expression of IRS-1 protein in liver by 245% in SHROB and 268% in SHR. Long-term inhibition of sympathetic activity with moxonidine therapy lowered free fatty acids and significantly improved insulin secretion, glucose disposal, and expression of key insulin signaling intermediates. Thus, moxonidine should be considered for the treatment of multiple metabolic and cardiovascular abnormalities associated with syndrome X.

Plasma Glucagon and Free Fatty Acid Responses to a Glucose Load in the Obese Spontaneous Hypertensive Rat (SHROB) Model of Metabolic Syndrome X

Metabolic Syndrome X is a cluster of abnormalities including insulin resistance, hyperlipidemia, hypertension, and obesity. We sought to determine if excess plasma glucagon and free fatty acids (FFA) might contribute to the insulin resistance in the obese spontaneous hypertensive rat (SHROB), a unique animal model of leptin resistance and metabolic Syndrome X. SHROB were extremely hyperinsulinemic and mildly glucose intolerant compared with lean SHR. SHROB had elevated fasting plasma glucagon and FFA, and showed paradoxical responses to an oral glucose challenge, with increased glucagon at 30 and 60 min postchallenge (200% ± 45% and 91% ± 13%, respectively; n = 9). In lean SHR, glucagon was nearly unchanged by glucose loading (<30% increase, P > 0.05; n = 5). Plasma FFA were not affected by a glucose load in SHROB, whereas SHR showed a decrease of 40% ± 6% (n = 5–9). The I/G molar ratio changed in opposite directions in the two genotypes, with a decrease in SHROB at 30 and 60 min, in contrast to the appropriate increase at 30 and 60 min postchallenge in the lean SHR (P < 0.01; n = 5–9). Administration of 500 ng/kg exogenous glucagon to SHR raised glucagon 56% ± 5% to a level that was similar to fasting SHROB. This level of circulating glucagon was sufficient to elevate glucose and insulin during the 7 hr of observation (n = 9). Based on these results, we suggest that fasting hyperglucagonemia and impaired suppression of glucagon secretion and FFA in response to an oral glucose load may contribute to insulin resistance and glucose intolerance in the SHROB model of metabolic Syndrome X.

Metabolic effects of antihypertensive agents: role of sympathoadrenal and renin-angiotensin systems

Reports of beneficial, neutral and adverse impacts of antihypertensive drug classes on glucose and lipid metabolism can be found in human data. Furthermore, mechanisms for these diverse effects are often speculative and controversial. Clinical trial data on the metabolic effects of antihypertensive agents are highly contradictory. Comparisons of clinical trials involving different agents are complicated by differences in the spectrum of metabolic disturbances that accompany hypertension in different groups of patients. Two physiological systems are predominant at the interface between metabolic and cardiovascular regulation: the sympathetic nervous system (SNS) and the renin-angiotensin system (RAS). These two systems are major targets of antihypertensive drug actions, and also mediate many of the beneficial and adverse effects of antihypertensive agents on glucose and lipid metabolism. Thiazides and β-adrenergic antagonists can adversely affect glucose and lipid metabolism, which are frequently compromised in human essential hypertension, and increase the incidence of new cases of diabetes. Laboratory studies confirm these effects, and suggest that compensatory activation of the SNS and RAS may be one mechanism. Other antihypertensives directly targeting the SNS and RAS may have beneficial effects on glucose and lipid metabolism, and may prevent diabetes. Resolution of the controversies surrounding the metabolic effects of antihypertensive agents can only be resolved by further laboratory studies, in addition to controlled clinical trials.