George R. Spratto

Ethanol suppression of naloxone-induced withdrawal in morphine-dependent rats

Abstract The technique of morphine pellet implantation was used to produce physical dependence on morphine in male rats. The number of “wet dog” shakes occurring within a period of 30 minutes during naloxone-precipitated (1.0 mg/kg, s.c.) withdrawal in four-day morphine implanted rats was determined after either acute or chronic treatment with ethanol. An acute dose of ethanol administered prior to withdrawal had no significant effect on the withdrawal response whereas chronic administration of ethanol during the development of dependence on morphine significantly suppressed the naloxone-precipitated withdrawal response to 44–57 percent of the control response. Analysis of brain and plasma for morphine concentration four days following dependence development showed no significant differences between morphine controls and those animals treated with both morphine and ethanol. Pentobarbital, another central nervous system depressant, demonstrated no effect on the withdrawal response, whether administered acutely or chronically during the development of dependence.

Locomotor activity in morphine-treated rats: Effects of and comparisons between cocaine, procaine, and lidocaine

The effects of cocaine, procaine, and lidocaine on open field and spontaneous (actophotometer) locomotor activities were assessed and compared in rats (1) treated acutely with morphine (single injection), (2) made dependent on morphine (SC pellets), (3) implanted with morphine and withdrawn at the time of peak dependence, and (4) implanted SC with lactose-containing pellets (sham). Cocaine-induced (10 or 30 mg/kg) open field and spontaneous locomotor activities were significantly greater in each of the four groups than those of the corresponding groups administered saline. Procaine (50 or 100 mg/kg) significantly reduced open field locomotor activity in all morphine-treated rats and spontaneous locomotor activity in acute rats. Lidocaine (30 mg/kg) significantly depressed spontaneous locomotor activity in acute rats. Upon comparison of the activities induced by the three local anesthetics, open field locomotor activity of sham-implanted rats was greater following cocaine (10 or 30 mg/kg) than following procaine (50 or 100 mg/kg). Only morphine withdrawn rats manifested greater activity following cocaine (10 mg/kg) than following either procaine (50 mg/kg) or lidocaine (10 mg/kg); activities were equivalent in dependent and acute rats. In contrast, cocaine-induced (30 mg/kg) open field locomotor activity of all morphine-treated rats was greater than either procaine- (100 mg/kg) or lidocaine- (30 mg/kg) induced activities. Spontaneous locomotor activity of all groups except acute morphine was greater following both doses of cocaine than following both doses of either procaine or lidocaine. In acute rats, only cocaine (10 mg/kg, induced greater activity than the other local anesthetics. Thus, stimulation of locomotor activity following cocaine treatment is a pharmacological property unique to cocaine and not shared by either procaine or lidocaine. Further, the data indicate that the methods selected for assessing locomotor activity may not give comparable results.

Body Temperature Response to Cocaine and Diazepam in Morphine-Treated Rats

Since cocaine or diazepam are used clinically and/or abused concomitantly with narcotics, this study was designed to determine if morphine-dependent, morphine-withdrawn, or acute morphine rats exhibit an altered core body temperature at 24 or 4 degrees C to either cocaine (10, 20, or 30 mg/kg i.p.) or diazepam (2 or 4 mg/kg i.p.). At 24 degrees C, each dose of cocaine manifested hyperthermia in all groups of rats except morphine-dependent whereas at 4 degrees C cocaine produced hypothermia in all groups except morphine-dependent. At both 24 and 4 degrees C, diazepam, 2 or 4 mg/kg, caused a hypothermic response in control, dependent, and withdrawn animals. At 24 degrees C, administration of acute morphine alone induced a hyperthermia which was antagonized by both doses of diazepam. At 4 degrees C, acute morphine alone induced an initial hypothermia followed by a hyperthermia; both doses of diazepam potentiated the hypothermic response to acute morphine. Thus, significant alterations in core body temperature may be induced following the administration of cocaine or diazepam to morphine-treated rats.

Brain and plasma concentrations of morphine during the development of physical dependence and tolerance

The subcutaneous implantation of a morphine pellet has been used to produce physical dependence and tolerance to morphine in mice (Maggiolo & Huidobro, 1961 ; Way, Loh & Shen, 1969) and rats (Cicero & Meyer, 1973). The degree of physical dependence and tolerance which develops in mice (Way & others, 1969) and rats (Cicero & Meyer, 1973) is comparable to that produced by an injection schedule of morphine. Recently, Patrick, Dewey & others (1975) reported that the concentration of morphine in the mouse brain increases rapidly and is maintained at a high level through the third day following morphine pellet implantation. This study was initiated in an effort to further quantify the technique of pellet implantation in the rat so that this model could be used for additional studies in our laboratory. Male, Sprague-Dawley derived rats (Laboratory Supply Co., Indianapolis, IN) six to eight weeks old (180-200 g) were employed. Food and water were freely available and animals were housed in community cages unless otherwise noted. Temperature was maintained between 21-23” and a 14 h light cycle (0600 to 2000 h) was employed. Two morphine pellets, each containing 103.5 mg morphine base and formulated according to the procedure of Gibson & Tingstad (1970), were implanted subcutaneously 2-4 cm posterior to the shoulders on the back of the animals under light ether anaesthesia. The mortality following implantation of two morphine pellets was between 3-5 % for all experiments provided that the animals were housed individually for at least 24 h following pellet implantation. Morphine pellets remained implanted in the rats during all experimental procedures except where noted. The degree of physical dependence on morphine was determined using a modification of the procedure of Cicero & Meyer (1973). Acute morphine withdrawal was precipitated by the administration of naloxone hydrochloride (0.4 mg kg-l, s.c.) and the number of ‘wet dog’ shakes were counted for a period of 30 min (a shorter period than that used by Cicero & Meyer but the results of the test are not affected). Symptoms of the withdrawal syndrome included writhing, salivation, lacrimation, teeth chattering, profuse diarrhoea, ejaculate-like discharge, hostility, bursts of activity which included escape behaviour and ‘wet dog’ shakes. Physical dependence, as determined by the number of ‘wet dog’ shakes, reached the highest level four to five days after implantation of 207 mg morphine base (ave. of 29 shakes per rat in 30 min on both days four and five). The number of ‘wet dog’ shakes per rat in 30 min

Potentiation of hexobarbital and amphetamine effects in male and female rats physically dependent on morphine.

Morphine dependence (following s.c. implantation of 207 mg morphine in pellet form) increases hexobarbital sleeping time in both male and female rats; this is accompanied by an apparent increase in brain sensitivity to hexobarbital. Only male morphine-dependent rats manifested a decrease in in vitro hepatic metabolism of hexobarbital as well as a significantly faster rate of decline of hexobarbital from the brain. In vivo uptake of hexobarbital into the brain revealed no difference between male and female morphine-dependent and sham-implanted animals. The suggested mechanism for the observed changes in male rats is a combination of inhibition of hepatic metabolism and an additive effect of morphine and hexobarbital. In female rats only an additive effect is manifested. In male rats morphine dependence also increases amphetamine-stimulated locomotor activity (open-field), increases amphetamine brain levels, and decreases in vitro metabolism of aniline. Morphine-dependent female rats manifested increases in brain levels of amphetamine but no other changes were observed. The in vivo decline of amphetamine from the brain of either male or female morphine-dependent rats was not significantly different from their respective controls. The suggested mechanism for the changes in male morphine-dependent rats is an increase in the brain uptake of amphetamine.

The comparative glycemic effects of catecholamines in rats and mice

Abstract Epinephrine, norepinephrine and isoproterenol were administered to either normally fed or 18 hr-fasted mice. While fasting did not alter the magnitude of epinephrine-induced hyperglycemia, it abolished norepinephrine-induced hyperglycemia. Isoproterenol produced hyperglycemia and depletion of liver glycogen in fed mice and hypoglycemia in fasted mice. In fed rats isoproterenol had no effect on blood glucose but elicited hyperglycemia in 24 hr-fasted rats. Isoproterenol also induced hyperglycemia in adrenalectomized normally fed mice. The results suggest that hyperglycemia in the fed mouse is a reflection of isoproterenol-induced glycogenolysis in the liver. It is suggested that the hypoglycemia in fasted mice is due to isoproterenol-stimulated insulin secretion. This study accents the criticality of the species employed as well as the pretreatment regimen with respect to data interpretation.

Failure of Beta-Adrenergic Blocking Agents to Antagonize the Hyperglycemic Effect of Epinephrine in Mice

Summary Chlorpromazine-induced hyperglycemia is abolished by adrenalectomy in mice. The administration of beta-adrenergic blocking agents failed to alter the magnitude of the CPZ-induced hyperglycemia. Similarly, beta-adrenergic blockade does not influence epinephrine-induced hyperglycemia in mice. Since beta-adrenolytic agents block both CPZ- and epinephrine-induced hyperglycemia in rats, the results of the present studies suggest a species difference in the susceptibility of certain adrenergic receptors to beta blockade.

Effects of the acetylcholinesterase inhibitor pinacolyl methylphosphonofluoridate (soman) on selected endocrine, glucose, and catecholamine levels in fasted and fed rats

Fed and 18-h fasted rats were given acute doses of either saline or the acetylcholinesterase inhibitor pinacolyl methylphosphonofluoridate (soman) 40, 60, or 80 micrograms/kg. After 30 min plasma samples were collected and assayed for glucose, insulin, glucagon, corticosterone, epinephrine and norepinephrine and the hypothalamus was isolated and assayed for acetylcholinesterase activity. Toxic sign scores were determined and they indicated that soman may be more toxic in the fasted rat. Soman-induced increases in corticosterone were observed in both fasted and fed rats; these levels were significantly higher in fasted rats given either soman or saline. Also, soman-induced increases in glucagon were more pronounced in fasted rats. Soman also caused an apparent dose-dependent increase in catecholamines and a decrease in hypothalamic acetylcholinesterase activity in both groups of rats. The expected lower insulin and glucose levels in the fasted rats were present in the saline-dosed animals and remained lower than fed rats after each dose of soman. This lack of soman-induced hyperglycemia may contribute to the toxicity of soman in fasted rats.

The effects of barbituric acid and 5-nitro barbituric acid on glycemic responses and in vitro insulin release in mice.

Abstract The effects of barbituric acid (BA) and 5-nitro BA on carbohydrate metabolism were assessed in mice. The administration of either BA or the 5-nitro analog elicited an initial hyperglycemia, glucose intolerance, and a decrease in responsiveness to tolbutamide. Although the glycemic responses produced by 5-nitro BA administration appeared greater than those elicited by the parent compound, these effects were abolished when the analog was administered to either fasted or adrenalectomized mice. In addition, 5-nitro BA did not inhibit d -glucose-stimulated insulin secretion in vitro . In contrast, BA attenuated both d -glucose- and tolbutamide-induced insulin release from isolated pancreatic islets. A competition for pancreatic β cell binding sites is offered as the possible mechanism of antagonism between BA and the sulfonylurea. The results indicate that while BA exerts its effects on carbohydrate metabolism directly on the pancreas, the effects of the 5-nitro analog appear to be mediated through stress-related factors. Because of its specificity for the insulin-secreting pancreatic β cells, structure-activity relationship studies with BA are likely to augment the development of a compound which will find clinical utility in the management of hyperinsulinism.

Examination of dihydrotachysterol-induced progeria as a model for aging changes in carbohydrate metabolism

Interest in the pharmacological effects of drugs in the elderly has created a need for a laboratory model in which responses of aged animals to drugs can be studied. Dihydrotachysterol (DHT)-induced progeria, a syndrome with symptoms similar to those seen in normal aging, was evaluated as an old age model. DHT-treatment was shown to produce a decreased carbohydrate tolerance that was due to an impaired release of insulin from pancreatic islets and not due to a decreased sensitivity to insulin. These changes were unlike those observed with aging. Thus, DHT-induced progeria would not appear to be a good model for aging for the investigation of carbohydrate metabolism. Evidence is presented which indicates that glucose and tolbutamide act via different mechanism to stimulate insulin release.