Ann Petro

Differential effects of fat and sucrose on the development of obesity and diabetes in C57BL/6J and A/J mice

We have previously demonstrated that the C57BL/6J (B/6J) mouse will develop severe obesity, hyperglycemia, and hyperinsulinemia if weaned onto a high-fat, high-sucrose (HH) diet. In the present study, we compared the effects of fat and sucrose separately and in combination on diabetes- and obesity-prone B/6J and diabetes- and obesity-resistant A/J mice. After 4 months, the feed efficiency ([FE] weight gained divided by calories consumed) did not differ across diets in A/J mice, but B/6J mice showed a significantly increased FE for fat. That is, B/6J mice gained more weight on high-fat diets without consuming more calories than A/J mice. The increase in FE was related to adipocyte hyperplasia in B/6J mice on high-fat diets. Fat-induced obesity in B/6J mice was unrelated to adrenal cortical activity. In the absence of fat, sucrose produced a decreased in FE in both strains. Animals fed a low-fat, high-sucrose (LH) diet were actually leaner than animals fed a high-complex-carbohydrate diet. Fat was also found to be the critical stimulus for hyperglycemia and hyperinsulinemia in B/6J mice. In the absence of fat, sucrose had no effect on plasma glucose or insulin. These data clearly show that across these two strains of mice, genetic differences in the metabolic response to fat are more important in the development of obesity and diabetes than the increased caloric content of a high-fat diet.

Strain-Specific Response toβ 3-Adrenergic Receptor Agonist Treatment of Diet-Induced Obesity in Mice1

Fat intake has long been associated with the development of obesity. The studies described herein show that fat adversely affects adipocyte adrenergic receptor (AR) expression and function. As beta 3AR agonists have been shown to acutely reduce adipose tissue mass and improve thermogenesis in genetically obese rodents, we examined whether chronic supplementation of a high fat diet with a highly selective beta 3AR agonist, CL316,243 could prevent diet-induced obesity, and whether the effect could be sustained over prolonged treatment. C57BL/6J and A/J mice were weaned onto one of three diets: low fat (10.5% calories from fat), high fat (58% calories from fat), or high fat supplemented with 0.001% CL316,243. B/6J mice gained more weight on the high fat diet than A/J mice (at 16 weeks: B/6J, 36.6 +/- 1.4 g; A/J, 32.9 +/- 0.8 g; P 0.05; n = 10). CL316,243 prevented the development of diet-induced obesity in A/J animals, but not in B/6J animals. A/J mice weighed 26.0 +/- 0.5 g at 16 weeks, whereas B/6J animals on the same diet weighed 34.1 +/- 0.8 g (P < 0.00001; n = 10), but food intake was not different between the strains throughout the study. beta-Adrenergic stimulation of adenylyl cyclase in obese B/6J mice was decreased by more than 75% in white adipose tissue and by more than 90% in brown adipose tissue (BAT). In contrast, in fat-fed A/J mice, beta-agonist-stimulated adenylyl cyclase was decreased in white adipose tissue by about 10%, whereas the activity in interscapular BAT was decreased by 50%, indicating significant retention of beta AR-stimulated activity in A/J mice compared to B/6J mice. High fat feeding was associated with decreased expression of beta 3AR and beta 1AR in white adipose tissue of both strains. However, chronic CL316,243 treatment prevented both the obesity and the decline in beta 3AR and beta 1AR messenger RNA levels in all adipose depots from A/J mice, but not B/6J mice. As CL316,243-treated A/J mice, but not B/6J mice, also showed marked uncoupling protein expression in white adipose depots, the ability of chronic CL316,243 treatment to prevent diet-induced obesity is dependent upon the elaboration of functional BAT in these regions.

Reversal of diet-induced obesity and diabetes in C57BL/6J mice.

We have previously shown that C57BL/6J (B6) mice develop severe obesity and diabetes if weaned onto high-fat diets, whereas A/J mice tend to be obesity and diabetes-resistant. The purpose of this study was to determine if obesity and diabetes in the B6 mouse could be completely reversed by reducing dietary fat content. After 4 months, both strains consumed more calories on a high-fat diet than on a low-fat diet, and both strains showed a higher feed efficiency (FE=weight gained/calories consumed) on the high-fat diet versus the low-fat diet. However, relative to A/J mice, B6 mice demonstrated a significantly higher FE on the high-fat diet. Hyperglycemia, hyperinsulinemia, and increased adiposity were apparent in B6 mice after 4 months on the high-fat diet regardless of whether the diet was begun at weaning or 4 months later. Correlational analyses showed that adiposity was strongly related to both insulin and glucose levels in B6 mice, but only moderately related to insulin levels in A/J mice. In obese B6 mice that were switched to a low-fat diet, obesity and diabetes were completely reversed. Adiposity, fasting glucose, and fasting insulin values in these mice were equivalent to those in B6 mice of the same age that had spent 8 months on the low-fat diet. In summary, our data show that in the B6 mouse the severity of diabetes is a direct function of obesity and diabetes is completely reversible by reducing dietary fat.

Low Plasma Leptin in Response to Dietary Fat in Diabetes- and Obesity-Prone Mice

Despite the fact that mutations resulting in the absence of leptin or its receptor have been associated with severe obesity and diabetes, such mutations do not appear to be responsible for most human obesity. Indeed, diet-induced obesity in animals and humans has been characterized by hyperleptinemia. This has been interpreted as evidence for leptin resistance. However, no careful longitudinal studies evaluating the role of leptin in the development of obesity exist. We report a series of studies in A/J and C57BL/6J (B/6) mice that demonstrate a direct relationship between the ability to increase plasma leptin levels in response to a high-fat diet and resistance to the subsequent development of obesity and diabetes. While leptin levels are similar in lean, low-fat-fed A/J and B/6 mice, the effects of a high-fat diet on plasma leptin differ dramatically between the two strains. After 4 weeks of high-fat feeding, leptin levels in A/J mice increased 10-fold, and this elevated level was maintained independent of weight gain throughout a 14-week feeding period. However, in B/6 mice, leptin levels remained at least twofold lower and only rose very gradually along with a significant increase in adiposity, hyperglycemia, and hyperinsulinemia. These differences in the response of leptin to diet are independent of food intake and plasma insulin levels during the 1st month of feeding. Further, we demonstrated that leptin administration did not influence the expression of the novel uncoupling protein UCP2, which also responds to dietary fat. From these results, we suggest that the response of leptin to fat feeding may be an important predictor of the development of subsequent obesity.

Nicotinic α7- or β2-containing receptor knockout: Effects on radial-arm maze learning and long-term nicotine consumption in mice

Classically, it has been thought that high-affinity nicotinic receptors-containing beta2 subunits are the most important receptor subtypes for nicotinic involvement in cognitive function and nicotine self-administration, while low affinity alpha7-containing nicotinic receptors have not been thought to be important. In the current study, we found that knockout of either beta2 or alpha7 subunits caused significant deficits in spatial discrimination in mice. The character of the impairment in the two knockouts was different. The beta2 knockout preferentially impaired cognition in males while the alpha7 caused impairment regardless of sex. Both beta2- and alpha7-containing nicotinic receptors also are important for nicotine self-administration, also in different ways. Most animal model studies of nicotine self-administration are relatively short-term whereas the problem of tobacco addiction is considerably longer-term. To better model the impact of nicotinic receptor subtypes on nicotine self-administration over the long-term, we studied the impact of genetic knockout of low affinity alpha7 receptors vs. high-affinity beta2-containing nicotinic receptors. Mice with knockouts of either of these receptors and their wildtype counter parts were given free access to a choice of nicotine-containing and nicotine-free solution in their home cages on a continuous basis over a period of 5 months. During the first few weeks, the beta2-containing nicotinic receptor knockout mice showed a significant decrease in nicotine consumption relative to wildtype mice, whereas the alpha7 knockout mice did not significantly differ from wildtype controls at the beginning of their access to nicotine. Interestingly, in the longer-term after the first few weeks of nicotine access, the beta2 knockout mice returned to wildtype mouse levels of nicotine consumption, whereas the alpha7 knockout mice developed an emergent decrease in nicotine consumption. The alpha7 receptor knockout-induced decrease in nicotine consumption persisted for the 5-month period of the study. Both alpha7- and beta2-containing nicotinic receptors play critical roles in cognitive function and nicotine self-administration. Regarding cognitive function, beta2-containing receptors are important for maintaining normal sex differences in spatial learning and memory, whereas alpha7 receptors are important for cognitive function regardless of sex. Regarding nicotine self-administration high-affinity beta2-containing nicotinic receptors are important for consumption during the initial phase of nicotine access, but it is the alpha7 nicotinic receptors that are important for the longer-term regulation of nicotine consumption.

Differential effects of fat and sucrose on body composition in A/J and C57BL/6 mice.

The C57BL/6 (B6) mouse is more sensitive to the effects of a high-fat diet than the A/J strain. The B6 mouse develops severe obesity, hyperglycemia, and hyperinsulinemia when fed this dietary regimen. This study was conducted to determine the effects of dietary fat and sucrose concentrations on body composition and intestinal sucrase (EC 3.2.1.48) and maltase (EC 3.2.1.20) activity in these two mouse strains. High-fat diets, regardless of sucrose content, resulted in significant weight gain, higher body fat, and lower body protein and water content in both strains of mice. The shift toward higher body fat and lower protein and water content was far greater in the B6 strain. Low-fat, high-sucrose diets resulted in lower body weight in both strains, as well as significantly greater body protein content in B6 mice. Analysis of intestinal sucrase showed that the enzyme was less active in B6 mice when the diet was high in sucrose. Both sucrase and maltase had lower activity in the presence of high dietary fat in both mouse strains. The percent reduction of intestinal enzyme activity due to dietary fat was similar in both strains. The B6 mouse exhibits disproportionate weight gain and altered body composition on a high-fat diet. This coupled with the reduced body weight and increased body protein on a low-fat, high-sucrose diet suggests that factors-relative to fat metabolism rather than sucrose metabolism are responsible for obesity.

Nicotine effects on learning in zebrafish: the role of dopaminergic systems

RationaleNicotine improves cognitive function in a number of animal models including rats, mice, monkeys, and recently, zebrafish. The zebrafish model allows higher throughput and ease in discovering mechanisms of cognitive improvement.Materials and methodsTo further characterize the neural bases of nicotine effects on learning in zebrafish, we determined changes in dopaminergic systems that accompany nicotine-enhanced learning.ResultsNicotine improved learning and increased brain levels of dihydroxyphenylacetic acid (DOPAC), the primary dopamine metabolite. There was a significant correlation between choice accuracy and DOPAC levels. The nicotinic antagonist mecamylamine blocked the nicotine-induced increase in DOPAC concentrations, in line with our previous finding that mecamylamine reversed nicotine-induced learning improvement.ConclusionsDopamine systems are related to learning in zebrafish; nicotine exposure increases both learning rates and DOPAC levels; and nicotinic antagonist administration blocks nicotine-induced rises in DOPAC concentrations. Rapid cognitive assessment of drugs with zebrafish could serve as a useful screening tool for the development of new therapeutics for cognitive dysfunction.

Increased nicotine self-administration following prenatal exposure in female rats

There is a significant association between maternal cigarette smoking during pregnancy and greater subsequent risk of smoking in female offspring. In animal models, prenatal nicotine exposure causes persistent alterations in cholinergic and monoaminergic systems, both of which are important for nicotine actions underlying tobacco addiction. Accordingly, the current study was conducted to determine if there is a cause-and-effect relationship between prenatal nicotine exposure and nicotine self-administration starting in adolescence. Pregnant rats were administered nicotine (6 mg/kg/day) by osmotic minipump infusion throughout gestation and then, beginning in adolescence and continuing into adulthood, female offspring were given access to nicotine via a standard operant IV self-administration procedure (0.03 mg/kg/infusion). Gestational nicotine exposure did not alter the initial rate of nicotine self-administration. However, when animals underwent one week of forced abstinence and then had a second opportunity to self-administer nicotine, the prenatally-exposed animals showed a significantly greater rate of self-administration than did the controls. Prenatal nicotine exposure causes increased nicotine self-administration, which is revealed only when the animals are allowed to experience a period of nicotine abstinence. This supports a cause-and-effect relationship between the higher rates of smoking in the daughters of women who smoke cigarettes during pregnancy and implicates a role for nicotine in this effect. Our results further characterize the long-term liabilities of maternal smoking but also point to the potential liabilities of nicotine-based treatments for smoking cessation during pregnancy.

Sazetidine-A, a Selective α4β2 Nicotinic Receptor Desensitizing Agent and Partial Agonist, Reduces Nicotine Self-Administration in Rats

Adequate treatment of tobacco addiction remains problematic. Part of the problem with treatment is a poor understanding of the pharmacologic aspects of nicotine contributing to addiction. In addition to activating nicotinic acetylcholine receptors, nicotine also desensitizes them. It is currently not known how much of each of nicotines actions contribute to its particular behavioral effects. Sazetidine-A (saz-A) is a novel nicotinic receptor-desensitizing agent and partial agonist with high selectivity for α4β2 receptors. The current experiments were conducted to determine whether saz-A would reduce nicotine self-administration in rats and to characterize its ancillary effects. Adult male Sprague-Dawley rats were allowed to self-administer nicotine. After initial food pellet training followed by 10 sessions of nicotine self-administration training, the rats were administered saz-A (0.1–3 mg/kg s.c.) or the saline vehicle in a repeated-measures counterbalanced design. Saz-A at the 3 mg/kg dose significantly decreased nicotine self-administration relative to performance of the same rats after saline injections. In a second study, long-term administration of this dose of sazetidine-A over the course of 10 sessions significantly reduced nicotine self-administration with no apparent diminution of effect. Saz-A in this dose range had only modest effects on locomotor activity, without any overall decrease in activity over a 1-h-long session. Saz-A significantly reduced food self-administration, but this effect was smaller than its effect on nicotine self-administration. Saz-A, which is a selective α4β2-desensitizing agent and partial agonist, effectively reduces nicotine self-administration. This type of treatment holds promise for a new therapy to aid smoking cessation.

Lorcaserin, a 5-HT2C Agonist, Decreases Nicotine Self-Administration in Female Rats

Lorcaserin, a selective 5-hydroxytryptamine2C (5-HT2C) agonist, has been shown to facilitate weight loss in obese populations. It was assessed for its efficacy in reducing nicotine self-administration in young adult female Sprague-Dawley rats. The effect of short-term doses (subcutaneous) on nicotine self-administration (0.03 mg/kg per infusion) with a fixed ratio 1 schedule was assessed in 3-h sessions. Short-term lorcaserin doses (0.3125–20 mg/kg) were administered in a counterbalanced order. Significant reduction of nicotine self-administration was achieved with all of the short-term doses in this range. Tests of lorcaserin on locomotor activity detected prominent sedative effects at doses greater than 1.25 mg/kg with more modest transient effects seen at 0.625 to 1.25 mg/kg. Long-term effects of lorcaserin on locomotor activity were tested with repeated injections with 0.625 mg/kg lorcaserin 10 times over 2 weeks. This low lorcaserin dose did not cause an overall change in locomotor activity relative to that of saline-injected controls. Long-term lorcaserin (0.625 mg/kg) significantly reduced nicotine self-administration over a 2-week period of repeated injections. Long-term lorcaserin at this same dose had no significant effects on food self-administration over the same 2-week period of repeated injections. These studies support development of the 5-HT2C agonist lorcaserin to aid tobacco smoking cessation.