David S. Roane

Distribution of glucokinase, glucose transporter GLUT2, sulfonylurea receptor-1, glucagon-like peptide-1 receptor and neuropeptide Y messenger RNAs in rat brain by quantitative real time RT-PCR

Glucokinase (GK), glucose transporter GLUT2, sulfonylurea receptor-1 (SUR1), glucagon-like peptide-1 receptor (GLP-1R) and neuropeptide Y (NPY) have been proposed to be involved in central glucose sensing or regulation of food intake. In this study, we combined tissue micropunch and real time reverse transcription polymerase chain reaction (RT-PCR), and measured GK, GLUT2, SUR1, GLP-1R and NPY mRNA expression in discrete areas in the hypothalamus and the hindbrain.

Synthesis of 2-piperidinecarboxylic acid derivatives as potential anticonvulsants

Abstract A variety of 2-piperidinecarboxamides were synthesized and evaluated for anticonvulsant activity using the MES and sc PTZ tests in mice and rats. Neurotoxicity was determined by the rotorod test. Several N-(benzyl)-2-piperidinecarboxamides exhibited potent MES activity in mice [2-CF3 14, ED50 = 29 mg/kg; 3-F 16, ED50 = 31 mg/kg; and 3-CF3 17, ED50 = 24 mg/kg]. The most active compounds in the MES test in mice were the 2,6-dimethylanilides [(R,S)-34, ED50 = 5.8 mg/kg; (R)-35, ED50 = 5.7 mg/kg; and (S)-36, ED50 = 14.8 mg/kg]. The enantiomer (S)-36 was about two-fold less potent in the MES test than (R)-35 and also was less neurotoxic. Acylation of the piperidine ring nitrogen of 12 and 34 led to a decrease in the MES activity. In the N-(α-methylbenzyl)-2-piperidinecarboxamides, the stereochemistry at either the 2-position of the piperidine ring or at the α-position of the N-(α-methylbenzyl) group does not significantly affect MES activity

Hydroxyl-substituted sulfonylureas as potent inhibitors of specific [3H]glyburide binding to rat brain synaptosomes.

We are seeking to discover potent CNS-active sulfonylureas with structural features that allow for the formation of several types of prodrugs. We report herein the syntheses of compounds comprising an initial series of hydroxyl-substituted analogues of the potent ATP-sensitive potassium channel blockers glyburide (glibenclamide) and gliquidone. Somewhat unexpectedly, several of the compounds were found to be comparably potent to glyburide as inhibitors of specific [(3)H]glyburide binding in rat brain preparations.

Short-term food restriction and refeeding alter expression of genes likely involved in brain glucosensing.

Several genes involved in glucosensing of the endocrine pancreas have been proposed to serve a similar function in the brain. These genes include the glucose transporter-2 (Glut-2) and glucokinase (GK). In addition, the glucagon-like peptide 1 receptor, which serves as a downstream signal modulator in pancreatic glucosensing and centrally alters feeding, is also of interest. We used quantitative real-time RT-PCR to measure changes in hypothalamic and brainstem Glut-2, GK, and Glp-1R expression of these genes induced by food restriction and refeeding. Sprague-Dawley rats were 50% food restricted for 1 day; one-half of the food-restricted rats were refed with chow for 1 hr before sacrifice. In both hypothalamus and brainstem, gene expression of Glut-2, GK, and Glp-1R was significantly lower in refed rats compared with food-restricted rats. The measures of gene expression in two feeding control groups (ad libitum and voluntarily overfed animals) were Intermediate between the food-restricted and refed groups, but were not significantly different from each other. The results indicate that putative glucosensing (GK, Glut-2, and Glp-1R) gene expression in the hypothalamus and brainstem is reduced in response to food Intake, depending on prior nutritional status.

Reduction of food intake and morphine analgesia by central glybenclamide

Previous research has indicated the presence of a reciprocal relationship between food intake and opioid-mediated analgesia. We believe the cellular candidate most likely acting as a common mediator of both ingestive and nociceptive behaviors is the ATP-sensitive K+ channel (K+ATP). This ion channel appears to be opened by mu and delta 1 opioid receptor agonists in the service of analgesia, and closed as cellular ATP availability rises. To further examine the role of the K+ATP in the relationship between feeding and opioid function, we administered 80 nmol of glybenclamide (a K+ATP antagonist) to male SD rats via the lateral ventricle. Chow consumption in the treated animals was significantly reduced over the following 48 h (F = 2.62, p < 0.013), with the peak effect (78% of control) occurring at 6 h. In the tail-flick test, 4 mg/kg morphine sulfate provided analgesia of 42.38 +/- 8.4% and 18.89 +/- 7.67% in vehicle and treated animals, respectively (p < 0.05, n = 8/group, one-tailed t-test). These results support the hypothesis that food intake and analgesia are reciprocally modulated through activity at the K+ATP.

Quinpirole-induced alterations of tail temperature appear as hyperalgesia in the radiant heat tail-flick test

Several reports in the recent literature argue both for and against the importance of alterations of tail-temperature in the outcome of the tail-flick test. The data we present here support the assertion that drug-induced changes of tail-temperature may have a highly significant effect on tail-flick latency independent of drug-induced changes in nociception. We previously reported that peripherally administered injections of the dopamine agonist, quinpirole, produce significant reductions in the latency of response in the tail-flick test. This present work confirms our earlier findings; however, it indicates that the apparent hyperalgesia is an artifactual function of quinpirole-induced increases in tail temperature. Quinpirole (0.1-1.0 mg/kg I.P.) produced significant (p < 0.001), dose-dependent, and highly correlated increases in tail temperature and decreases in tail-flick latency 15 min following injection. When controls for the change in tail temperature were applied, there was no distinguishable effect of the drug on tail-flick latencies. Sixty minutes following the administration of quinpirole there was no observable effect of the drug on either tail-temperature or tail-flick latency. The results of this study indicate that a) peripherally administered quinpirole has no effect on nociception as measured in the tail-flick test apart from its ability to alter tail temperature; and b) alterations in tail temperature may significantly alter the outcome of the tail-flick test.

Expression Levels of Genes Likely Involved in Glucose-sensing in the Obese Zucker Rat Brain

Abstract It has been suggested that certain cells in the brain, like pancreatic β-cells, use glucose transporter-2 (GLUT-2), glucokinase and glucagon-like peptide-1 receptor (GLP-1R) to sense glucose in the service of multiple aspects of energy balance. The obese Zucker rat displays numerous disturbances in energy homeostasis and may provide a model of dysfunctional expression of genes related to nutrient control systems. Using real-time RT-PCR we measured gene expression for three of the pancreatic glucose-sensing markers and neuropeptide Y (NPY) in the medial, lateral hypothalamus and hindbrain of lean and obese Zucker rats of both genders. Additionally, we measured circulating levels of glucose, leptin, insulin, corticosterone and glucagon. The results indicate that GLUT-2 mRNA expression is decreased, whereas glucokinase is increased in the hindbrain of obese rats. NPY mRNA level is significantly higher, whereas GLP-1R is significantly lower in the medial hypothalamus in obese individuals. Gender-related differences were found in the hindbrain and medial hypothalamus for GLUT-2 and in the lateral hypothalamus for GLP-1R and they may be related to the fact that the female Zucker rats do not develop diabetes as readily as males. Furthermore, the hindbrain may be an important site for glucose-sensing where major phenotypic changes occur for glucose-sensing genes expression.

Effect of Additives on the Diffusion of Ketoprofen Through Human Skin

AbstractThe influence of Carbopol® polymer, types C-934P and C-940, and the penetration enhancer oleic acid on transdermal permeation of ketoprofen through a fullthickness human skin was investigated. Ketoprofen patches were fabricated; and permeation parameters such as the flux. permeability coeficient. enhancement ratio, lag time, and partition coefficients were determined. The results indicated a maximum flux of 7.778 μg/cm2/hrfrom the patches made with C-934P when the oleic acid concentration was 35%. The enhancement ratio was 22.8. The maximum flux value for patches made from C-940 was obtained with 10% oleic acid. The corresponding enhancement ratio was 34.25. The results indicated that the concentration of oleic acid needed for maximum flux depends upon the type of Carbopol polymer selected for the study. Further, ketoprofen transdermal drug delivery systems can be fabricated to obtain a zero-order release through human skin.

Evidence of hyperglycemic hyperalgesia by quinpirole

Male albino rats were tested for antinociception following injections (IP) with saline, quinpirole (Quin) (1 mg/kg), morphine sulfate (M.S.) (5 mg/kg), or both Quin and M.S. (1 mg/kg and 5 mg/kg, respectively). Quin reduced and M.S. increased tail-flick latency as compared to controls. Tail-flick latencies of the animals injected with both drugs were significantly reduced as compared M.S. alone. Quin increased blood glucose levels by 96 percent, as compared to saline controls. In competitive binding studies Quin displaced 3H-DAGO (IC50 = 29.8 microM). CD-1 mice demonstrated a naloxone-reversible analgesia following ICV Quin (100 micrograms). These data are consistent with the hypothesis that the hyperglycemic effects of Quin attenuate M.S. analgesia while the antinociceptive effects of Quin may be mediated through opioid receptors.

The Effect of Centrally Administered Glibenclamide, Tolbutamide and Diazoxide on Feeding in Rats.

While there are many theories on the control of feeding behavior that emphasize a role for energy substrates and their metabolism, the mechanism that couples changes in energy substrate supply and metabolism to alterations in food intake remains unclear. The purpose of the present project was to investigate the possibility that central ATP-sensitive potassium (KATP(+)) channels may serve as integrators between cellular energetics and alterations in neuronal activity that control feeding, such that pharmacologic manipulation of the channels would result in alterations in feeding behavior. Intracerebroventricular (ICV) injections of the KATP(+) channel blocker glibenclamide significantly increased feeding in fasted and fed male Sprague-Dawley rats. Likewise, the first generation sulfonylurea, tolbutamide, also increased feeding. ICV injection of the KATP(+) channel opener, diazoxide, modestly inhibited feeding. These results suggest that central KATP(+) channels may be involved in the regulation of feeding behavior.