Donald E. Hutchings

The ontogeny of cannabinoid receptors in the brain of postnatal and aging rats

It is recognized that a number of the biological effects of delta 9-tetrahydrocannabinol (THC) can be attributed to a cannabinoid receptor found in abundance in the brain. Due to observations that cannabinoid drugs exert some developmental toxicity, it was of interest to examine the developmental pattern of cannabinoid receptors in the brain of neonatal rats through young adulthood, and then to further examine the cannabinoid receptor during the aging process in the brain of rats 3 to 32 months of age. Using radioligand binding assays, this study demonstrated that cannabinoid receptor binding capacity increases progressively from birth to postnatal day (PND) 60. Within the striatum, a significant increase in binding occurred between PNDs 14 and 21. In the cerebellum, cannabinoid receptor binding capacity doubled at 7-day postnatal intervals until adulthood. Cannabinoid receptor binding in the cortex doubled between PNDs 7 and 14. Within the hippocampus, there were small incremental increases until the final adult level was reached at PND 21. There was no significant alteration in the affinity for CP-55940 during development. These findings might reflect an increased differentiation of neurons into cells possessing cannabinoid receptors, or an increase in the number of cannabinoid receptors on cell bodies or projections in regions undergoing developmental changes. Once the adult cannabinoid receptor levels have been reached, binding activity in the whole brain preparation neither increased nor declined during the normal aging process.

Plasma concentrations of delta-9-tetrahydrocannabinol in dams and fetuses following acute or multiple prenatal dosing in rats☆

Delta-9-tetrahydrocannabinol (THC) was administered by gastric intubation to pregnant rats to study the effects of dose-level and dosing regimen on plasma concentration in dams and fetuses. Two multiple-dose groups were administered either 15 or 50 mg/kg of delta-9-THC once daily during the last two weeks of gestation. Two acute groups were administered the same dose as above but only once on the last day of gestation. Sixty min after receiving the last dose all dams and their fetuses were sacrificed by decapitation, blood collected, centrifuged and plasma removed. Quantitative measurement of delta-9-THC in plasma was carried out using GS/MS. Among the dams, plasma concentrations covaried with dose and multiple dosing produced higher concentrations than acute, especially at the high dose. Among the fetuses, plasma concentrations were approximately 10% of those found in the dams. The fetuses from the high, multiple-dose dams similarly yielded significantly higher concentrations. These findings are discussed with respect to other studies of the placental transfer of delta-9-THC and effects of postnatal developmental.

The puzzle of cocaine's effects following maternal use during pregnancy: are there reconcilable differences?

This is a selective review of the clinical and epidemiological literature. It attempts to reconcile disparate and contradictory findings dealing with the morphologic, growth, and neurobehavioral effects reported to occur in neonates and young children exposed prenatally to cocaine. A history of cocaine use in the United States is briefly presented followed by impressionistic observations of some of the events that transpired during the cocaine epidemic of the 1980s. Based on the collective research findings, it is tentatively suggested that the teratogenic effects of prenatal cocaine may be produced only in those infants exposed to the highest doses reported in the literature. It remains unknown, however, whether or not these effects may be dependent on the concurrent abuse of alcohol and/or cigarettes. As to growth and neurobehavioral outcomes, effects attributable primarily to cocaine alone and not other substances of abuse appear to be only marginal and transitory. The data to support these conclusions are particularly tenuous and are thus offered only as working hypotheses. Because of the intractable methodological and interpretive problems inherent in human developmental research on substance abuse, any attempt to draw definitive conclusion is admittedly premature. Because these methodological problems also complicate the efforts of ongoing studies, answers to these persistent questions may not be readily forthcoming.

Comparison of oral and subcutaneous routes of cocaine administration on behavior, plasma drug concentration and toxicity in female rats

Oral and subcutaneous routes of administration of cocaine HCl were investigated in female Wistar rats for food and water consumption, locomotor activity, stereotypic behaviors, plasma drug concentrations and injection site pathology. Animals received either 40 or 80 mg/kg/day by gastric intubation (PO-40 and PO-80 respectively) or 20 or 40 mg/kg/day subcutaneously (SC-20 and SC-40). All groups received the drug or the vehicle for 16 consecutive days. Locomotor activity and stereotypy were evaluated on Days 1, 5, 10, and 15. Plasma drug concentrations and injection site pathology were determined on Day 16. Subcutaneous administration was associated with a sensitization to the effects of cocaine on locomotion and stereotypy, higher blood levels than oral administration at the same dose, and severe dermal lesions. However, there were no differences in any measure between the SC-20 and SC-40 groups. Oral cocaine was also associated with behavioral sensitization. However, unlike the SC route, oral cocaine was characterized by dose-related increases in locomotion and stereotypy in the absence of gastrointestinal pathology. Inasmuch as oral administration resulted in dose-response relationships and low toxicity while subcutaneous administration did not, these factors should be considered in future studies utilizing chronic cocaine administration.

Issues of risk assessment : lessons from the use and abuse of drugs during pregnancy

Therapeutic uses and risks associated with the opioids, cannabis, cocaine, and phencyclidine are discussed as well as limitations of the developmental animal and clinical pediatric literature, especially with respect to problems of quantitative risk assessment. Human and animal developmental findings for methadone and cannabis are compared with respect to long-term behavioral effects with special emphasis on pharmacological and interpretive issues. It is suggested that neonatal withdrawal is the most serious neurobehavioral sequela associated with maternal use of opioid and other sedative-hypnotic compounds. Withdrawal phenomena are described and attempts to develop animal models are discussed. Methodological considerations including surrogate fostering, pair-feeding, and problems associated with the administration of compounds to lactating dams are discussed in the context of the adequacy of the EPA developmental neurotoxicity battery to characterize risk for abuse substances as well as the new NIDA medication compounds.

Delta-9-tetrahydrocannabinol during pregnancy in the rat: I. Differential effects on maternal nutrition, embryotoxicity, and growth in the offspring☆

Either 15 or 50 mg/kg of delta-9-tetrahydrocannabinol (THC) in sesame oil was administered by gastric intubation to gravid rats during the last two weeks of gestation. A pair-fed control group was administered the vehicle alone and allowed to eat and drink only the amount consumed by the 50 mg/kg group on the same gestation days. A nontreated control group was left undisturbed during pregnancy. All treated and control litters were fostered at birth to untreated dams. Among the dams receiving 50 mg/kg of THC, food and water intake was initially reduced to 75-80% of nontreated controls but then recovered over 3-4 days to approximately a 15-20% reduction until term. Compared with the nontreated dams, both THC dose-level groups and pair-fed control dams gained significantly less body weight from conception to term. Offspring mortality did not differ between the nontreated and pair-fed controls but was significantly higher among both dose-level THC exposed groups. In addition, there was a dose-related increase in the sex-ratio of live male to female offspring as well as significant effects on rate of growth for both sexes. The results are discussed with respect to published animal and clinical studies of cannabinoid exposure during pregnancy.

Neonatal Withdrawal Following Pre- and Postnatal Exposure to Methadone in the Rat

Recent evidence has shown that infant rats undergo precipitated withdrawal following chronic postnatal injection of morphine. In this study we examined whether or not infants exposed to methadone prenatally via the placental blood supply and postnatally via the dams milk would also experience precipitated withdrawal. Dams were implanted on gestational day 14 with osmotic minipumps containing one of two concentrations of methadone to supply the opiate throughout gestation and the first postnatal week. Nontreated and pair-fed controls were used. On postnatal day 7, pups were injected with naltrexone and their locomotor activity and ultrasonic vocalizations measured. Methadone exposed pups were more active and vocalized more when injected with naltrexone than with saline. The controls did not show these behavioral changes. The milk of methadone-exposed dams apparently contains sufficient quantities of the opiate for dependence to develop. The results are consistent with other data that demonstrate that very young rat pups can experience an opiate abstinence syndrome that includes increased behavioral activation.

Delta-9-tetrahydrocannabinol during pregnancy in the rat: II. Effects on ontogeny of locomotor activity and nipple attachment in the offspring

Either 15 or 50 mg/kg delta-9-tetrahydrocannabinol (THC) in sesame oil was administered by gastric intubation to gravid rats during the last two weeks of gestation. A pair-fed control group was administered the vehicle alone and allowed to eat and drink only the amount consumed by the 50 mg/kg group on the same gestation days. A nontreated control group was left undisturbed during pregnancy. All treated and control litters were fostered at birth to untreated dams. Intact litters from the two THC treated and the two control groups were tested at 3-day intervals from birth to 32 days of age for differences in locomotor activity. In addition, pups were tested for nipple attachment on days 2, 5, 8, 11, and 14 of age. There were no differences in locomotor activity among any of the groups although activity level varied during development. Pups from dams exposed to 50 mg/kg of THC, as well as the pair-fed controls, displayed significantly longer latencies to attach to a nipple. These results suggest that the impaired nipple attachment observed among the high-dose offspring was not a primary effect of THC, but rather was secondary to the significant reduction of food and water intake among the dams. The behavioral findings are discussed with respect to other animal and clinical reports of prenatal cannabinoid exposure.

Prenatal administration of buprenorphine using the osmotic minipump: a preliminary study of maternal and offspring toxicity and growth in the rat

Buprenorphine, an opioid with mixed agonist-antagonist properties, is gaining new attention as an effective pharmacotherapy for opioid and possibly cocaine abuse. With a view to its consideration for use with pregnant clients and because so little is know of its potential developmental toxicity, we have carried out this preliminary study. Three doses of buprenorphine (BUP) were administered by osmotic minipump from day 8 of gestation through parturition. In addition to 0.3, 1.0, and 3.0 mg/kg/day of BUP, a vehicle control group received sterile water via minipump and a nontreated control group was left undisturbed during pregnancy. All treated and control litters were fostered at birth to untreated dams. BUP produced a dose response reduction in maternal water intake but had no effect on maternal weight gain, the frequency of resorptions, or birthweight. BUP had no effect on perinatal mortality and produced inconsistent effects on postnatal growth. The unique chemical and pharmacological properties of this compound, especially its bell-shaped or asymptotic dose response effects, are discussed with respect to the development of an adequate animal model to evaluate neurobehavioral effects and assess its safety for use during pregnancy.

Prenatal administration of methadone using the osmotic minipump: Effects on maternal and offspring toxicity, growth, and behavior in the rat

Two doses of methadone were administered by osmotic minipump from day 8 of gestation through parturition. A pair-fed control group received saline via minipump and was allowed to eat and drink only the amount consumed by the high dose group on the same gestation days. A nontreated control group was left undisturbed during pregnancy. All treated and control litters were fostered at birth to untreated dams. Naloxone challenge of the dams after parturition showed that drug treatment produced physical dependence. Methadone treatment reduced maternal weight gain but had no effect on either the frequency of resorptions or birthweight. Both doses of methadone increased perinatal mortality but only the high dose produced a decrement in postnatal growth. To examine the effects of methadone on the rest-activity cycle of the offspring, groups of three littermates from each of the treated and control groups were tested for an 8 h observation period on electronic activity monitors at 22 days of age. No behavioral effects were observed for either control group or the low dose methadone group. The high dose methadone offspring, however, spent less time resting, showed disrupted rhythmicity, and poor state regulation. These findings are discussed in relation to earlier studies using once per day methadone administration as well as clinical descriptions of infants undergoing opiate abstinence.