Prenatal inhibition of hypothalamic sex steroid uptake by cocaine: effects on neurobehavioral sexual differentiation in male rats

Several adrenergically active drugs have been shown to prevent the masculinizing and/or defeminizing effects of testosterone on brain sexual differentiation. We examined the ability of the neuronal norepinephrine uptake blocker, cocaine, to produce similar effects. The ability of cocaine to inhibit sex steroid incorporation into the hypothalamus during a critical period for sexual differentiation of the brain was examined in females treated at birth with testosterone. Sixty minutes after administration, cocaine was observed to inhibit both testosterone and estradiol incorporation into the hypothalamus by approximately 50%. Long-term consequences of prenatal cocaine exposure were studied by injecting Sprague-Dawley dams twice daily with 3, 10 or 30 mg/kg of cocaine hydrochloride on days 15 through 20 of gestation and examining the offspring. In adulthood, cocaine-exposed males, but not females were found to exhibit significantly less marking behavior than controls. Cocaine-exposed males in the 10 mg/kg group tested for sex behavior exhibited demasculinization in some aspects of the behaviors tested. Measurement of plasma hormone levels in this group revealed elevated levels of plasma LH, but normal levels of FSH and testosterone. No differences were observed in cocaine-exposed males with respect to sex organ or adrenal weights, but thymus was approximately 25% smaller compared to control males at 80 days of age. In a separate experiment, dams were treated with 3 mg/kg of cocaine twice daily from days 15 through 21 of gestation and half of the male pups received additional injections twice a day for the first 5 days postnatally.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prenatal cocaine alters later responses to morphine in adult male mice

Mice prenatally exposed to cocaine (25 mg/kg), physiological saline or non-treated during the last 6 days of pregnancy were evaluated as adults for the rewarding properties of 2 mg/kg of morphine, using the conditioned place preference (CPP) procedure. Likewise, isolated animals underwent a social interaction test with conspecifics after receiving the same morphine dose. Unlike control or animals pre-treated with saline, subjects prenatally treated with cocaine did not develop CPP with this dose of morphine. Only cocaine-exposed animals showed increased threat, avoidance and fleeing during the social encounter. No differences in motor effects of morphine were observed. Analysis of monoamines revealed effects of housing conditions, isolated animals having fewer DOPAC but higher levels of HVA than those grouped, but in both groups there was a decrease in DOPAC in cocaine- and saline-treated mice. Prenatal cocaine exposure decreases the response to the rewarding properties of drugs in mature offspring. They also implicate cocaine consumption during pregnancy could affect the response of offspring to take other drugs of abuse.     

Prenatal cocaine exposure decreases brain-derived neurotrophic factor proteins in the rat brain

The pregnant rats received daily sc injections of cocaine (30 mg/kg) or saline from the gestational day (GD) 7 to GD 20. At 1 week postnatal, all pups were killed and the hippocampus, cortex and striatum were dissected out. Levels of brain-derived neurotrophic factor (BDNF) under the basal condition and depolarization with high potassium (40 mM) were measured. The results showed that hippocampal BDNF levels under basal and depolarization conditions were all significantly lower in the pups prenatally exposed to cocaine than those exposed to saline. There were no significant differences in basal BDNF levels between the cocaine and saline groups in the cortex or striatum. However, the prenatally cocaine-treated pups showed significantly less BDNF release following high potassium depolarization than the saline-treated animals did in both these regions. The results support the suggestion that prenatal cocaine exposure decreases BDNF expression in the offspring.     

Prenatal cocaine exposure potentiates 5-HT(2a) receptor function in male and female rat offspring

We have reported previously prenatal cocaine-induced functional deficits in serotonergic terminals, and gender-specific supersensitivity of postsynaptic 5-HT(1A) receptor-mediated hormone responses in offspring. This study investigates the effects of prenatal exposure to cocaine on postsynaptic 5-HT(2A) receptor-mediated responses in prepubescent male and female offspring. Pregnant rats were administered saline or (-)cocaine (15 mg/kg, s.c., b.i.d) from gestational day 13 through 20. Changes in 5-HT(2A) receptor function in offspring were assessed by differences in the ability of DOI [4-iodo, 2,5-dimethoxyphenyl-isopropylamine; 2. 0 mg/kg, s.c.] to elevate plasma levels of the hormones ACTH, corticosterone and renin. Basal hormone levels in male and female progeny were unaffected by prenatal cocaine exposure. However, prenatal exposure to cocaine significantly potentiated the magnitude of the ACTH response to DOI in both male (+19%) and female (+43%) progeny. Similarly, the DOI-induced elevation of plasma renin was markedly potentiated in male (+51%) and female (+83%) cocaine-exposed offspring. Although DOI significantly elevated corticosterone levels in both male and female offspring, the magnitude of corticosterone responses was not altered by prenatal exposure to cocaine. Densities of agonist ((125)I-DOI)-labeled receptors in hypothalamus and cortex were unaltered by prenatal exposure to cocaine. These data indicate prenatal cocaine-induced supersensitivity of postsynaptic 5-HT(2A) receptor function in male and female offspring without changes in receptor density. Synapse: 35:163-172, 2000.     

Exposure to cadmium during gestation and lactation decreases cocaine self-administration in rats

This investigation examined the effects of perinatal cadmium exposure on subsequent self-administration of cocaine during the adult cycle. Female Sprague-Dawley rats were gavaged daily with 0.0 (14% sucrose solution, w/v) or 5.0 mg cadmium chloride (dissolved in 14% sucrose solution, w/v) for 30 days prior to breeding with non-exposed males. Dams continued to experience cadmium exposure through gestation and until pups were weaned at postnatal day (PND) 21. On PND 70, offspring were anesthetized and chronic indwelling jugular catheters were implanted. Following recovery, test subjects were trained in operant chambers to self-administer 0.500 mg/kg infusion (inf) intravenous cocaine on a fixed-ratio (FR) 2 schedule of reinforcement. Following acquisition, self-administration rates were tested for saline, 0.030, 0.060, 0.125, 0.250, and 0.500 mg/kg inf cocaine. Rats exposed developmentally to cadmium self-administered significantly less than controls at saline, 0.030, and 0.060 mg/kg inf cocaine. These data indicate that early-life cadmium exposure, a common exposure vector of which is the use of tobacco products, may affect cocaine sensitivity.     

Somatostatin receptors increase in the olfactory bulb of developing pups after perinatal exposure to cocaine

The effects of chronic prenatal and/or postnatal exposure to cocaine on somatostatin concentration and receptors were studied in the olfactory bulbs of rat pups at birth and at 15 days old. Wistar rats were injected subcutaneously with single daily doses of 40 mg cocaine hydrochloride/kg from days 7 to 19 of gestation, from day 7 of gestation to day 15 postpartum or from parturation to day 15 postpartum. Fetal exposure to cocaine decreased SS concentrations in the olfactory bulb of the newborn pups while prenatal-plus-postnatal exposure increased this parameter. Administration of cocaine only during lactation did not induce any change. Exposure during gestation or during nursing induced an increase in the total number of somatostatin receptors and a decrease in the affinity constant in the olfactory bulb of newborn and 15-day-old pups. These results suggest that the development of somatostatin receptors in the olfactory bulb can be altered by prenatal and/or nursing period exposure to cocaine.     

D-cycloserine facilitates extinction of cocaine self-administration in rats

Previous research has indicated that D-cycloserine [DCS; a N-methyl-D-aspartate (NMDA) partial agonist] facilitates the extinction of conditioned fear as well as the extinction of cocaine conditioned place preference. Sprague Dawley rats were first trained to self-administer cocaine and then we compared their extinction behavior (lever pressing) following treatment with vehicle; 15 mg/kg DCS; or 30 mg/kg DCS. We showed that 30 mg/kg DCS, but not 15 mg/kg significantly accelerated extinction of cocaine self-administration behavior when compared with saline by almost half (4 days vs. 9 days). At 2 weeks when all animals had extinguished, there were no longer differences between the groups. The present findings support of the potential of NMDA partial agonists as prospectively valuable in facilitating the extinction of cocaine-seeking behavior. More specifically, we demonstrate that 30 mg/kg DCS was effective at significantly accelerating the extinction of cocaine self-administration behavior in rats. These results provide further support for the potential of DCS as a treatment strategy for addiction.     

Long-term consequences of prenatal exposure to cocaine or related drugs: Effects on rat brain monoaminergic receptors

Reports from both this laboratory and others indicate that prenatal exposure of rats to cocaine can produce alterations in development, activity and responses to environmental stimuli. In order to determine a biochemical basis for these effects, radioligand receptor-binding assays for different monoaminergic receptors were performed on rat brain tissues obtained from offspring of dams treated SC with saline, cocaine (15 mg/kg b.i.d.), amitriptyline (10 mg/kg) or amfonelic acid (AFA, 1.5 mg/kg). Male rat pups were fostered by surrogate dams and one rat per litter taken at 30, 60 or 180 days postnatal for determination of striatal and prefrontal cortical D2 receptors, prefrontal cortical 5HT2 receptors, cortical alpha 1-, alpha 2-, beta 1- and beta 2-adrenoceptors. Across all drug treatments and times, the only significant change was at 30 days of age when beta 1-adrenoceptors were increased 68% in the cocaine exposed pups--a time when these rats show hyperactivity--and at 180 days postnatal when a 20% decrease in DA2 receptor Bmax was observed. Also, cortical membrane Mg(2+)-dependent Na+, K(+)-ATPase activities and basal ATPase activities were unaltered by any of the treatments at any of the times. These results suggest that few changes have occurred in monoaminergic receptor sensitivity as a result of the exposure to these drugs during gestation. The behavioral changes that are known to occur following prenatal exposure to cocaine may be due to presynaptic alterations in neurotransmitter function rather than changes in postsynaptic receptors.     

Developmental effects of prenatal cocaine exposure on 5-HT1A receptors in male and female rat offspring

Prenatal cocaine exposure results in behavioral abnormalities throughout development in rats, but little is known regarding the biological mechanisms underlying these abnormalities. Pregnant rats received subcutaneous twice-daily injections (1 ml/kg) of normal saline or 15 mg/kg of cocaine hydrochloride throughout gestation (gestation days 1-20). Following delivery, pups were placed with untreated surrogates. Male and female pups were killed on postnatal days 30, 60 or 120 for assessment of 5-HT(1A) receptor development in the forebrain, diencephalon, midbrain and pons using radiolabel immunocytochemistry. Findings revealed gender and age differences in developmental regulation of 5-HT(1A) receptors, indicating that male rats are more susceptible to long-term consequences of prenatal cocaine exposure in comparison to females. This study also demonstrates gender-specific development of serotonin (5-HT(1A)) receptors across postnatal ages, demonstrating a fundamentally different pattern of development of 5-HT(1A) receptors between males and females.     

Downregulation of kappa-opioid receptors in basolateral amygdala and septum of rats withdrawn for 14 days from an escalating dose "binge" cocaine administration paradigm

There is evidence showing that the opioid systems play an important role in cocaine addiction; fewer studies have examined their roles in cocaine withdrawal. This study was conducted to determine whether cocaine or chronic withdrawal from cocaine alters the receptor component of the kappa-opioid system. Male Fischer rats were injected with saline or cocaine (3x15 mg/kg/day for 4 days, 3x20 mg/kg/day for 4 days, 3x25 mg/kg/day for 4 days, and 3x30 mg/kg/day for 2 days), three times daily at 1-h intervals in an escalating dose paradigm for 14 days. Identically treated rats were withdrawn from cocaine or saline for 14 days. We performed quantitative autoradiographic mapping of kappa-opioid receptors (KOP-r) in the brains of rats treated with this escalating dose "binge" cocaine administration paradigm and of rats withdrawn from cocaine for 14 days. A significant condition (chronic/withdrawal) effect was shown across all regions analyzed. A significant increase in [3H]CI-977 binding to KOP-r was detected in the septum of rats treated with an escalating dose binge cocaine administration paradigm and killed 30 min after the last cocaine injection. In contrast, there was a decrease in KOP-r binding in the septum and the basolateral amygdala of rats withdrawn for 14 days from chronic escalating dose binge cocaine administration, compared to rats at the end of 14 days chronic escalating dose cocaine administration. These results reconfirm and extend that KOP-r undergoes upregulation in response to chronic binge cocaine administration here, with an escalating dose. The observed lowering in KOP-r binding, which was shown in two brain regions of cocaine withdrawn animals, might contribute to the persistent dysphoria reported a long time after the discontinuation of the drug.     

Neurobehavioral toxicity of triethyltin in rats as a function of age at postnatal exposure

Triethyltin (TET) has been shown to be neurotoxic when injected on postnatal day (PND) 5. In the present experiment we examined the toxicity of a single exposure to TET at several postnatal ages. Rat pups were injected ip with 0 (saline), 1.5, 3.0, or 6.0 mg/kg TET bromide on PND 1, 5, 10 or 15. In agreement with our previous data, PND-5 exposure to 6 mg/kg TET produced behavioral toxicity and decreased adult brain weight. High dose pups were less successful in descending on a rope at 20 and 21 days of age, and were hyperactive in figure-eight mazes at 29-30 and 57-58 days of age. The spatial distribution of activity was also altered: photocell counts were increased primarily in the figure-eight area of the maze. The size of the milk bands was reduced in 6 mg/kg pups injected on either PND 1 or PND 5. Preweaning growth was decreased following all injection ages; this reduction was most pronounced for pups exposed to TET on PND 1 and PND 5. Mating behavior was disrupted in 6 mg/kg males irrespective of age at exposure. These data demonstrate a differential sensitivity to the toxicity of TET during postnatal life, with maximal susceptibility on PND 5.     

Prenatal cocaine alters dopamine transporter binding in postnatal day 10 rat striatum

The effect of in utero cocaine exposure on the postnatal binding of the radiolabeled dopamine (DA) uptake inhibitor [₃H]GBR 12935 to the DA uptake complex was examined in male rats. One set of pregnant Sprague-Dawley rats was given subcutaneous (s.c.) injections of cocaine (40 mg/kg) or 0.9% saline from gestational day (GD) 8–21. Another set of animals received bilateral s.c. Silastic implants, each containing 60 mg cocaine base dissolved in polyethylene glycol (PEG) or PEG only, from GD 18-21. The density of[₃H]GBR 12935 binding to the DA transporter in striatum and mesencephalon was assessed by quantitative autoradiography on postnatal day (PND) 1, 10, 30, and 60. Both treatment methods resulted in a decrease of [₃H]GBR 12935 binding in dorsal lateral striatum of cocaine-exposed offspring on PND 10. There were no significant differences in [₃H]GBR 12935 binding between offspring of cocaine and vehicle-treated dams at any other time points examined. Thus, prenatal cocaine exposure by either daily injection from GD 8–21 or continuous infusion from GD 18–21 resulted in a transient decrease in DA transporter binding in the dorsal lateral striatum that was apparent on PND 10. © 1996 Wiley-Liss, Inc.     

Effects of chronic cocaine administration on aggressive behavior in virgin rats

Virgin Sprague-Dawley rats received subcutaneous injections of saline, 3.5, 7.5 or 15 mg/kg of cocaine, twice daily, for 20 consecutive days. Females were videotaped for 10 minutes in the presence of a male rat for assessment of aggression towards the intruder 2, 3, and 5 days following cessation of cocaine or saline administration. Oxytocin levels in discrete brain areas were assayed following behavioral testing, 5 days following cessation of cocaine or saline administration. The 30 mg/kg-dose group tended to have a lower frequency of fight attacks and aggressive postures compared to saline-treated controls across sessions. The frequency of most of the behaviors analyzed were represented by quadratic functions across time, such that the highest frequency of behavior occurred 2 days following the final injection with relatively less activity 3 and 5 days following cessation of saline or cocaine administration. The 30 mg/kg cocaine-treated group had significantly lower hippocampal OT levels than the 15 mg/kg group 5 days following cessation of cocaine or saline administration.     

Reduced serotonin release and serotonin uptake sites in the rat nucleus accumbens and striatum after prenatal cocaine exposure

This study was designed to assess the effects of prenatal cocaine exposure on the development of the serotonergic system. Pregnant Sprague-Dawley rats received daily sc injections of either cocaine (30 mg/kg) or saline from gestation day 7 (GD 7) to GD 20. At 1 week postnatal, all pups were killed and tissues containing the striatum and nucleus accumbens dissected out. In superfusion experiments, tissue slices were incubated with [3H]serotonin ([3H]5-HT) for 30 min and then superfused. The [3H]5-HT release was induced by exposures to the following treatments: electrical stimulations (20 mA or 40 mA, 0.5 Hz, 4 min), the medium containing 15 or 30 mM potassium (2 min), fenfluramine (1 or 2 microM for 2 min), para-chloroamphetamine (1 or 2 microM for 2 min), methiothepin (1 or 2 microM for 2 min), and fluoxetine (1 or 2 microM for 2 min). The results showed that the treatment-induced [3H]5-HT releases were all significantly less pronounced in the pups prenatally exposed to cocaine than in those prenatally exposed to saline regardless of the mechanisms by which the treatment increases extracellular 5-HT. Saturation analysis showed that the Bmax of [3H]citalopram binding sites was also significantly lower in the pups prenatally treated with cocaine than in those prenatally treated with saline. The results are consistent with the concept that less serotonergic innervation may exist in the examined brain areas of cocaine-treated offspring at 1 week postnatal, and support the hypothesis that prenatal cocaine exposure affects the postnatal development of the serotonergic system.     

Primates exposed to cocaine in utero display reduced density and number of cerebral cortical neurons.

This study examined the effects of cocaine use during the second trimester of pregnancy on cerebral neocortical volume and density, and total number of neocortical neurons and glia in offspring. We also evaluated the extent of postnatal recovery of cytoarchitectural abnormalities previously observed in the neocortex of two-month-old primates born from cocaine-treated mothers (Lidow [1995] Synapse 21:332-334). Pregnant monkeys received cocaine orally (20 mg/kg/day) from the 40th to 102nd days of pregnancy (embryonic day [E]40-E102). On E64 and E65, the animals were injected with [(3)H]thymidine. Cerebral hemispheres of the offspring were examined at three years of age. We found a reduction in the neocortical volume and density and total number of neocortical neurons. The observed reduction in neuronal number within the neocortex was not accounted for by the increase in the number of neurons in the white matter of cocaine-exposed animals, because the number of these "extra" neurons was equal to only half that of missing neurons. We detected no significant changes in the number of neocortical glia. The cytoarchitectural abnormalities in the neocortex of prenatally cocaine-exposed three-year-old monkeys closely resembled previously described neocortical abnormalities in similarly exposed two-month-old animals: the neocortex lacked a discernible lamination; the majority of the cells labeled by [(3)H]thymidine injected during neocortical neurogenesis did not reach their proper position within the cortical plate. Therefore, postnatal maturation is not associated with significant improvement in neocortical organization in primates prenatally exposed to cocaine. There was, however, a postnatal recovery of low glial fibrillary acidic protein (GFAP) immunoreactivity previously observed in 2-month-old cocaine-exposed animals.     

Consequences of maternal cocaine on cerebral microvascular functions in piglets

Maternal cocaine abuse is associated with fetal and neonatal neurological abnormalities. Prolonged exposure to cocaine can induce blood flow disorders, growth restriction, and hypoxia in the newborn. We investigated the impact of chronic fetal cocaine exposure on cerebral microvascular reactivity and autonomic function in the piglets. Pregnant pigs received cocaine (1 mg/kg i.v.; twice weekly) or saline throughout the last trimester. Prenatal exposure to cocaine did not have any significant effect on the birth weight of the piglets as compared to the control. Following delivery, effects of recurrent prenatal cocaine exposure on cerebral microvascular functions were examined in piglets (3-6 days old). Pial arteriolar responses to applications of 5-hydroxytryptamine (5-HT), endothelin-1 (ET-1), and clonidine were examined using closed cranial windows. Functional effects of prenatal cocaine exposure on changes in mean arterial pressure (MAP) and pial arteriolar diameter induced by intracisternal injection (i.c.) of clonidine (1 microg/kg) were also determined. Topical applications of 5-HT, ET-1, and clonidine dose-dependently decreased pial arteriolar diameter in the control and these constrictions were significantly enhanced in the in utero cocaine-exposed piglets. Prenatal cocaine exposure did not have any significant effects on the resting MAP and heart rate as there were no differences between the groups. IC clonidine caused sustained decrease in MAP in both groups but the decrease was more pronounced in the cocaine than the control group. IC clonidine causes cerebral microvascular dilation coincident with the development of hypotension. Such dilation was severely attenuated in the cocaine group, even though the hypotension was much more pronounced than in the control. In conclusion, prenatal cocaine exposure resulted in attenuated autoregulatory vasodilation and potentiated responses to vasoconstrictor agents. The mechanisms behind the effects of in utero cocaine exposure on alteration of newborn cerebral functions need further investigation. Such actions may be important in development of cerebral pathologies associated with recurrent prenatal cocaine exposure.     

Inhibition of morphine withdrawal by the NMDA receptor antagonist MK-801 in rat is age-dependent

This study investigated the effects of the NMDA receptor antagonist MK-801 on the development of morphine dependence in 7-, 14-, and 21-day-old rat pups. For 6.5 days, starting at 1, 8, or 15 days of age, rats were pretreated with MK-801 (0.03 or 0.1 mg/kg, bid) or saline; 15 min later, morphine sulfate (10 mg/kg) or saline was injected to induce opiate dependence. On the afternoon of the seventh day, pups were injected with MK-801 (0.1 mg/kg) or saline and 15 min later with naltrexone (1 mg/kg) to precipitate withdrawal. Pups were then placed in a warm chamber with the litter and their behavior scan-sampled every 15 sec for a total of 15 min. MK-801 failed to inhibit morphine withdrawal in the 7-day-old rat, but did attenuate the development of morphine dependence in both the 14- and 21-day-old rats. These results suggest that the NMDA receptor is not functionally active in opiate withdrawal until around the second to third week of postnatal life in the rat and that there exists a transition period for the NMDA receptor to play a role in the development of opiate dependence and withdrawal.     

Altered cocaine-induced behavioral sensitization in adult mice exposed to cocaine in utero

Behavioral sensitization induced by psychostimulants is characterized by increased locomotion and stereotypy and may reflect aspects of neuronal adaptations underlying drug addiction in humans. To study the developmental contributions to addictive behaviors, we measured behavioral responses in adult offspring to a cocaine sensitization paradigm following prenatal cocaine exposure. Pregnant Swiss-Webster (SW) mice were injected twice daily from embryonic days 8 to 17 (E8-E17, inclusive) with cocaine (20 or 40 mg/kg/day; COC20 and COC40, respectively), or saline vehicle (SAL and SPF40) subcutaneously (s.c.). A nutritional control group of dams were 'pair-fed' with COC40 dams (SPF40). P120 male offspring from each prenatal treatment group were assigned to a behavioral sensitization group and injected with cocaine (15 mg/kg) or saline intraperitoneally (i.p.) every other day for seven doses. Locomotor activity and stereotypy were measured during habituation, cocaine initiation, and following a cocaine challenge 21 days after the last initiation injection. As expected, animals demonstrated significantly more locomotion and stereotypic behavior following acute and recurrent injection of cocaine compared to saline-injected animals. However, for each prenatal treatment group, cocaine-sensitized animals showed unique temporal profiles for the increase in locomotor sensitization and stereotypy over the course of the sensitization protocol. Two features that distinguished the altered behavioral progression of prenatally cocaine-exposed animals (COC40) from control (SAL) animals included blunted augmentation of locomotion and enhanced patterns of stereotypic behavior. These findings provide evidence that the behavioral activating effects of cocaine in adult animals are altered following exposure to cocaine in utero.