Sonya K. Sobrian

Prenatal cocaine and/or nicotine exposure produces depression and anxiety in aging rats.

The adult use of cocaine and nicotine has been linked to depression and/or anxiety. Changes in emotional behavior were assessed using behavioral paradigms developed as animal analogs of psychiatric disorders in 12-14 month old Sprague-Dawley rats exposed daily on gestational days 8-20 to cocaine and nicotine, either alone or in combination. Results from the elevated plus maze (EPM), used to assess anxiety-related behaviors, indicated that offspring prenatally exposed to either high-dose cocaine (40 mg/kg/day) or high-dose nicotine (5.0 mg/kg/day) were less timid/more impulsive. Animals from these two groups spent the most time on the open arms, and had the highest percentage of entries into the open arms of the EPM. Combined in utero exposure to cocaine and nicotine nullified these effects. Cocaine challenge (20 mg/kg) did not interact with prenatal treatment, but increased activity on all arms of the EPM in all groups. Sucrose preference was used as a measure of anhedonia, a cardinal symptom of depressive illness. Reduced sucrose preference was seen only in the group of offspring prenatally exposed to high-dose cocaine (40 mg/kg) plus low-dose nicotine (2.5 mg/kg/day). Exposure to a water-deprivation stress normalized sucrose preference in this group, without altering preference or intake in the other prenatal treatment groups. Transient hyperactivity was seen in the offspring of dams treated with high-dose nicotine, an effect that was again reversed in combined drug groups. Traditional gender differences in activity levels and sucrose intake, that is, females greater than males, were still evident in this population of aging rats. These data indicate that prenatal exposure to cocaine and/or nicotine has long-term effects on emotional behavior. Combined drug exposure contributed to the development of depressive symptoms, but not anxiety-like behavior, in a dose-dependent manner. In contrast, exposure to high doses of either drug alone reduced cautionary behavior. Data from this line of research could provide insight into the pathogenesis of emotional disorders, especially during the aging process.

Influence of prenatal maternal stress on the immunocompetence of the offspring.

To evaluate the effects of prenatal maternal stress on the development of humoral immunocompetence in the offspring and on their hormonal and immunologic responses to postnatal stress, gravid Sprague-Dawley rats were exposed daily on gestational days 15-21 to prenatal environmental stress [(PES) 15 unsignaled, inescapable electric foot-shocks (0.05 mA for 0.5 s)] or prenatal psychological stress [(PPS) pregnant rats were placed in the nonelectrified section of the apparatus and allowed to see, hear, and smell a nonpregnant partner being environmentally stressed]. Pregnant controls (PC) were placed in the apparatus for 30 min. Serum corticosterone (CCS) and immunoglobulin G (IgG) levels were measured in the offspring every 7 days from birth to postnatal day (PND) 28. On PND 29-33, offspring were environmentally stressed; hormonal and immune status were determined on PND 34. Levels of IgG were reduced in PES and PPS offspring on PND 0 and in PES offspring on PND 7 and 28. These changes were unrelated to differences in CCS and did not reflect altered maternal-pup interactions or nutritional factors. Postnatal stress was immunosuppressive in PC pups but did not alter immune parameters in PPS offspring. In PES females, postnatal stress was also immunosuppressive. However, in PES males with already reduced IgG levels postnatal stress enhanced immune function. These data provide the first experimental evidence that prenatal maternal stress can alter immune parameters in the rat offspring.

Interactive effects of prenatal cocaine and nicotine exposure on maternal toxicity, postnatal development and behavior in the rat.

Two experiments were performed to investigate the interactive effects of prenatal coadministration of cocaine hydrochloride (C) and nicotine tartrate (N). Experiment I was designed to determine doses of C and N that could be coadministered without altering maternal gestational parameters and/or fetal viability. Exposure of Sprague-Dawley rats to combined high-dose C (20 mg/kg) and high-dose N (5.0 mg/kg) on gestation days 8–21 was not more toxic to dam or fetus than that of exposure to C alone. Experiment II investigated pregnancy outcome, postnatal development, and behavior of the offspring following drug exposure to either high-dose cocaine (20 mg/kg: CS), high-dose nicotine (5.0 mg/kg: NS), or both (NC) on gestation days 8–21. N was administered by osmotic minipump and C by sc injection. Saline-injected dams, fitted with saline-filled pumps (SS), and untreated dams, pair-fed (PF) to NC females, served as controls. Alterations in maternal variables were limited to a 10–15% decrease in food consumption in NC and CS groups. Pregnancy outcome and birth statistics were unaffected by prenatal treatment, as was offspring body weight during the first four postnatal weeks. However, the development of surface righting was delayed in CS pups, and only CS offspring were underresponsive to the stimulatory effects of dopamine agonists on activity and stereotypy. Behavioral responses to N challenge were similar in all groups. In addition, only CS offspring showed altered behavioral responses in a spontaneous alternation task. Treatment effects on dopamine D1 and D2 binding in the caudate nucleus were not observed. The combination of N and C did not exacerbate any of the behavioral changes seen in CS offspring. These results support the hypothesis that C is a behavioral teratogen in rodents, and suggest that in the present model, nicotine can mitigate some of the consequences ofin utero exposure to cocaine.

Recommendations concerning the new U.S. National Institutes of Health initiative to balance the sex of cells and animals in preclinical research

The U.S. National Institutes of Health (NIH) announced last May that steps will be taken to address the over‐reliance on male cells and animals in preclinical research. To further address this announcement, in September 2014, scientists with varying perspectives came together at Georgetown University to discuss the following questions. (1) What metrics should the NIH use to assess tangible progress on policy changes designed to address the over‐reliance on male cells and animals in preclinical research? (2) How effective can education be in reducing the over‐reliance on male cells and animals in preclinical research and what educational initiatives sponsored by the NIH would most likely effect change? (3) What criteria should the NIH use to determine rigorously defined exceptions to the future proposal requirement of a balance of male and female cells and animals in preclinical studies? (4) What additional strategies in addition to proposal requirements should NIH use to reduce the overreliance of male cells and animals in preclinical research? The resulting consensus presented herein includes input from researchers not only from diverse disciplines of basic and translational science including biology, cell and molecular biology, biochemistry, physiology, pharmacology, neuroscience, cardiology, endocrinology, nephrology, psychiatry, and obstetrics and gynecology, but also from recognized experts in publishing, industry, advocacy, science policy, clinical medicine, and population health. We offer our recommendations to aid the NIH as it selects, implements, monitors, and optimizes strategies to correct the over‐reliance on male cells and animals in preclinical research.—Sandberg, K., Umans, J. G., Georgetown Consensus Conference Work Group. Recommendations concerning the new U.S. National Institutes of Health initiative to balance the sex of cells and animals in preclinical research. FASEB J. 29, 1646‐1652 (2015). www.fasebj.org

Prenatal antiepileptic drug exposure alters seizure susceptibility in rats

An animal model is used to address the issue of prenatal exposure to certain antiepileptic drugs and seizure susceptibility in the offspring. Administration of doses established as median therapeutic doses in humans of phenobarbital, valproate and clonazepam to pregnant rats during the last third of gestation produced sexually dimorphic alterations in pentylenetetrazol (PTZ)-induced seizures as well as in non-convulsive (spontaneous alternation and cliff avoidance) behaviors in the offspring. Altered seizure susceptibility occurred in the absence of overtly recognizable morphological abnormalities and did not appear to reflect differences in the status of circulating drug-binding plasma proteins. Possible neural and/or metabolic mechanisms responsible for these behavioral changes are discussed.

Determining normal variability in a developmental neurotoxicity test A report from the ILSI Research Foundation/Risk Science Institute expert working group on neurodevelopmental endpoints

With the implementation of the Food Quality Protection Act in 1996, more detailed evaluations of possible health effects of pesticides on developing organisms have been required. As a result, considerable developmental neurotoxicity (DNT) data have been generated on a variety of endpoints, including developmental changes in motor activity, auditory startle habituation, and various learning and memory parameters. One issue in interpreting these data is the level of variability for the measures used in these studies: excessive variability can obscure treatment-related effects, or conversely, small but statistically significant changes could be viewed as treatment related, when they might in fact be within the normal range. To aid laboratories in designing useful DNT studies for regulatory consideration, an operational framework for evaluating observed variability in study data has been developed. Elements of the framework suggest how an investigator might approach characterization of variability in the dataset; identification of appropriate datasets for comparison; evaluation of similarities and differences in variability between these datasets, and of possible sources of the variability, including those related to test conduct and test design. A case study using auditory startle habituation data is then presented, employing the elements of this proposed approach.

Behavioral response profiles following drug challenge with dopamine receptor subtype agonists and antagonists in developing rat.

As part of an investigation into the effects of gestational ethanol (ETOH) exposure on the developing dopamine (DA) system, pregnant Sprague-Dawley rats were exposed to one of three conditions: ETOH, pair-fed (PF) to the ETOH group, or ad libitum lab chow controls (LC). In this paper we report behavioral drug challenge effects for offspring of the two control groups (PF and LC). Male and female pups between postnatal days (PNDs) 21 and 23 in age were exposed to one of three intraperitoneal/subcutaneous doses of one of eight drugs chosen to assess the functional status of the DA D(1), D(2), and D(3) receptor subtype, or a saline control. Agonists were SKF 38393, apomorphine (APO), quinpirole (QUIN), and 7-hydroxy-N,N-di-n-propyl-2-amino-tetralin [7-OH-DPAT (DPAT)]; antagonists were spiperone (SPIP), SCH 23390, and two recently developed D(3) antagonists nafadotride (NAF) and PD 152255. Immediately following drug injection, pups were placed in observation cages, where eight behaviors (square entries, grooming, circling, rearing, sniffing, head and oral movements, and yawning) were scored at 3-min intervals for 30 min. Classic behavioral profiles were generally obtained for the high-dose mixed agonists APO, DPAT, and QUIN, which potently increased square entries, rearing, and sniffing, while reducing grooming and head movements. However, low-dose APO had no effect on behavior. The D(1) agonist, SKF 38393, had a strikingly different behavioral profile; it had no effect on square entries at any dose, while increasing grooming and sniffing at the medium dose. The D(1) antagonist, SCH 23390, profoundly decreased all behaviors except oral and head movements, especially at high doses. In contrast, the effects of the D(2) antagonist, SPIP, were limited to increasing sniffing at the medium dose. The two putative D(3) antagonists, NAF and PD 152255, presented strikingly different profiles. NAF induced a pattern of behavioral suppression that resembled the profile of high-dose SCH, while high-dose PD 152255 stimulated behavior. The failure of low-dose APO to have any effect on behavior suggests that the D(2) autoreceptor is not functional in preweanling rats. This hypothesis is further supported by the lack of behavioral suppression seen with low-dose QUIN and DPAT. Failure of NAF to produce behavioral activation at low doses and the stimulatory effects seen with PD 152255 suggests that either the D(3) autoreceptor, the postsynaptic D(3) receptor, or both are not fully functional at this age as well.

Social behavior of offspring following prenatal cocaine exposure in rodents: a comparison with prenatal alcohol

Clinical and experimental reports suggest that prenatal cocaine exposure (PCE) alters the offsprings’ social interactions with caregivers and conspecifics. Children exposed to prenatal cocaine show deficits in caregiver attachment and play behavior. In animal models, a developmental pattern of effects that range from deficits in play and social interaction during adolescence, to aggressive reactions during competition in adulthood is seen. This review will focus primarily on the effects of PCE on social behaviors involving conspecifics in animal models. Social relationships are critical to the developing organism; maternally directed interactions are necessary for initial survival. Juvenile rats deprived of play behavior, one of the earliest forms of non-mother directed social behaviors in rodents, show deficits in learning tasks and sexual competence. Social behavior is inherently complex. Because the emergence of appropriate social skills involves the interplay between various conceptual and biological facets of behavior and social information, it may be a particularly sensitive measure of prenatal insult. The social behavior surveyed include social interactions, play behavior/fighting, scent marking, and aggressive behavior in the offspring, as well as aspects of maternal behavior. The goal is to determine if there is a consensus of results in the literature with respect to PCE and social behaviors, and to discuss discrepant findings in terms of exposure models, the paradigms, and dependent variables, as well as housing conditions, and the sex and age of the offspring at testing. As there is increasing evidence that deficits in social behavior may be sequelae of developmental exposure alcohol, we compare changes in social behaviors reported for prenatal alcohol with those reported for prenatal cocaine. Shortcomings in the both literatures are identified and addressed in an effort to improve the translational value of future experimentation.

Effects of pre- and neonatal nicotine exposure in rodents: inconsistent evidence.

This review presents an analysis of the literature on behavioral effects of developmental exposure to nicotine, as assessed in rodent models that mimic the consequences for human offspring of maternal cigarette smoking. Despite the frequency of reports of low birth weight, hyperactivity, cognitive deficits, and psychiatric problems, inconsistencies exist in both the clinical and experimental literature. Confounding socioeconomic and other demographic variables may account for discrepancies in clinical reports, and the choice of developmental exposure period and the method of nicotine administration may explain differences in experimental outcomes. Analysis of a number of variables (e.g., physical, behavioral, and cognitive) shows that fetal exposure to nicotine does not consistently cause growth retardation or decreased birth weight, nor reliably affect motor activity. But combined pre- and neonatal exposure is likely to result in delayed reflex development, global impairments in learning and memory, and an increased incidence of symptoms that model psychiatric illness. There is also support for increased self-administration of nicotine and other drugs of abuse in animals exposed developmentally to nicotine, as well as potent effects on offspring responses to drug challenges. Unlike reports in the clinical literature, sexually dimorphic effects were not evident in most animal models. Possible neuroanatomical and cholinergic mechanisms responsible for behavioral changes are briefly discussed. Statistical and design considerations are provided to increase the translational value of this research and, most importantly, enhance the replicability of reported findings.

Prenatal Nicotine and/or Cocaine Differentially Alters Nicotine-Induced Sensitization in Aging Offspring

Repeated exposure to psychostimulant drugs can result in behavioral sensitization, an amplified response in locomotor activity and stereotypy, which is used to model aspects of drug addiction. The expression of behavioral sensitization, induced by i.p. injections of nicotine once daily for 5 days, was examined in 450‐day‐old male rats exposed prenatally on GD 8–20 to one of the following conditions: (1) low nicotine: 2.5 mg/kg/day nicotine [LN]; (2) high nicotine: 5.0 mg/kg/day nicotine [HN]; (3) low nicotine/high cocaine: 2.5 mg/kg/day nicotine plus 40 mg/kg/day cocaine [LN/HC]; (4) high nicotine/low cocaine: 5.0 mg/kg/day nicotine plus 20 mg/kg/day cocaine [HN/LC]; (5) pair‐fed controls: food intake yoked to HC dams [PF]; and (6) saline controls: daily injections of 0.9% NaCl solution[SAL]. Initial injection of nicotine did not alter activity or stereotypy in comparison to saline injections, with offspring in all prenatal treatment groups showing a desensitization to nicotine. Five consecutive daily nicotine injections resulted in behavioral sensitization in HN and HN/LC prenatal drug groups. Offspring exhibited an increase in horizontal activity that was evident on day 3, and still present after a 1.0 mg/kg i.p. nicotine challenge 72 hours after the last injection (day 8). SAL offspring exhibited attenuated sensitization. In contrast, nicotine sensitization was not seen in the LN, HC/LN, and the PF offspring; activity remained at the level seen after the initial injection of nicotine. Moreover, nicotine significantly reduced total activity in the LN and PF groups in comparison with their saline‐injected counterparts. These data suggest that gestational exposure to high‐dose nicotine, either alone or in combination with cocaine, may carry a greater risk than low‐dose nicotine exposure of stimulant abuse in later life.