This is your teen brain on drugs: In search of biological factors unique to dependence toxicity in adolescence.

Response variability across the lifespan is an important consideration in toxicology and risk assessment, and the toxic effects of drugs and chemicals during adolescence need more research. This paper summarizes a workshop presented in March 2019, at the Society of Toxicology Annual Meeting in Baltimore, Maryland, that brought together experts in research on drug dependence and toxicity related to nicotine, cannabis, cocaine, and other illicit drugs during adolescence. The goal of the workshop was to address the following issues: (1) Do the effects of adolescent exposure differ from the same exposure in adults? (2) Are there unique biological markers of adolescent brain development? If so, what are they and how reliable are they? (3) Since multiple factors influence substance use disorder, can we disentangle risk factors for abuse and/or toxicity? What are the underlying biological susceptibilities that lead to dependence and neurotoxicity? What are the social, psychosocial and environmental factors that contribute to abuse susceptibilities? This paper reviews drug policy and national trends in adolescent substance use; the public health consequences of e-cigarettes; rat models of adolescent-onset nicotine self-administration and persisting effects of gestational nicotine; sex-dependent effects of delta-9-tetrahydrocannabinol on adolescent brain-behavior relationships; and translational approaches for identifying adolescent risk factors for transition to drug dependence. There is strong evidence that drug exposure prior to adulthood has longer lasting effects on behavior and the underlying neural circuitry. These effects, which are sex-dependent and influenced by stress, may be candidates as predictors of adolescent vulnerability. A major challenge to determining if adolescents have a unique susceptibility to dependence is whether and to what extent the human data allow distinction between the increased risk due to biological immaturity, an underlying biological susceptibility to dependence, or psychosocial and environmental factors for substance dependence. Factors important to consider for development of animal models include the timing and pattern of exposure as it relates to adolescence; age of assessment, and direct comparison with similar effects following exposures to adults to demonstrate that these effects are unique to adolescence. Here we provide a roadmap for further research into what makes adolescent brain development unique.

Single trial nicotine conditioned place preference in pre-adolescent male and female rats.

The mean age of first voluntary tobacco inhalation is 12.3 years (DiFranza et al., 2004). 60% of smokers start smoking before the age of 14 and 90% are dependent before reaching the age of 19. Females are typically more sensitive to nicotine than males yet few studies examine the effects of nicotine on the reward systems in pre-adolescent female subjects. This study utilized the single trial conditioned place preference (CPP) test in very young (postnatal day 25-27) rats of both sexes. Latent effects on anxiety and amphetamine response were determined 5 and 7 days following a second nicotine exposure. Results show that 0.05 mg/kg nicotine induced CPP in females following a single trial while both sexes showed CPP following the 0.5 mg/kg dose. Five days later, rats dosed with 0.05 mg/kg show increased time on the open arm of the elevated plus maze, an anxiolytic response. While baseline activity was increased in nicotine-exposed males 7 days following dosing, amphetamine response was not affected by the treatments in either sex. Therefore, our data suggest that young females are more sensitive to nicotine reward than males supporting a heightened sensitivity of the mesolimbic dopamine system in very young females. However, alterations in baseline activity were only seen in males suggesting that different components of the system are affected by nicotine in each sex. An anxiolytic response to nicotine 5 days after dosing may suggest that this very young age group is uniquely affected by this very low nicotine dose. Clearly, nicotine has substantial acute and lasting effects during pre-adolescence at doses substantially lower than seen at older ages as reported by others. These effects, which could potentially result from cigarette or e-cigarette smoking by 11-12 year old children , focus attention on the vulnerability of this age group to nicotine.

Pretreatment with Δ9-tetrahydrocannabinol (THC) increases cocaine-stimulated activity in adolescent but not adult male rats.

Marijuana (Cannabis sativa) remains one of the most widely used illegal drugs, with adolescents being particularly vulnerable to its use and abuse. In spite of this, most studies are conducted in adult animals even though the effects might be quite different in adolescents. Additionally, the use of marijuana often precedes the use of other psychoactive drugs including cocaine, especially when marijuana exposure begins during early adolescence. The purpose of this study was to examine the effects of repeated Δ9-tetrahydrocannabinol (THC), the major active ingredient in marijuana, in adolescents compared to adults and to determine its subsequent effects on cocaine-stimulated activity. To this end, adolescent (postnatal day PND 34) and adult (PND 66) rats were administered 3 mg/kg/day THC for 8 days and locomotor activity was measured on days 1, 2, 7 and 8 after dosing. On day 12 (4 days after the last dose of THC), rats were injected with escalating doses of cocaine and behavior was recorded. Results show that THC depressed locomotor activity in adult rats but not in adolescents. However, following a cocaine challenge, adolescents exposed to THC showed increased locomotor responses to cocaine compared to chronic vehicle-injected controls. This was not seen in adults. These results show that the effects of cocaine are enhanced after THC in adolescents, but not adults, and that this might account for the greater transition to cocaine after early, as opposed to later, marijuana use.

Maternal cannabis use alters ventral striatal dopamine D2 gene regulation in the offspring.

BACKGROUND: Prenatal cannabis exposure has been linked to addiction vulnerability, but the neurobiology underlying this risk is unknown. METHODS: Striatal dopamine and opioid-related genes were studied in human fetal subjects exposed to cannabis (as well as cigarettes and alcohol). Cannabis-related gene disturbances observed in the human fetus were subsequently characterized with an animal model of prenatal Δ-9-tetrahydrocannabinol (THC) (.15 mg/kg) exposure. RESULTS: Prenatal cannabis exposure decreased dopamine receptor D2 (DRD2) messenger RNA expression in the human ventral striatum (nucleus accumbens [NAc]), a key brain reward region. No significant alterations were observed for the other genes in cannabis-exposed subjects. Maternal cigarette use was associated with reduced NAc prodynorphin messenger RNA expression, and alcohol exposure induced broad alterations primarily in the dorsal striatum of most genes. To explore the mechanisms underlying the cannabis-associated disturbances, we exposed pregnant rats to THC and examined the epigenetic regulation of the NAc Drd2 gene in their offspring at postnatal day 2, comparable to the human fetal period studied, and in adulthood. Chromatin immunoprecipitation of the adult NAc revealed increased 2meH3K9 repressive mark and decreased 3meH3K4 and RNA polymerase II at the Drd2 gene locus in the THC-exposed offspring. Decreased Drd2 expression was accompanied by reduced dopamine D2 receptor (D(2)R) binding sites and increased sensitivity to opiate reward in adulthood. CONCLUSIONS: These data suggest that maternal cannabis use alters developmental regulation of mesolimbic D(2)R in offspring through epigenetic mechanisms that regulate histone lysine methylation, and the ensuing reduction of D(2)R might contribute to addiction vulnerability later in life.

Methylphenidate response in prenatal cocaine-exposed rats: a behavioral and brain functional study.

Prenatal cocaine exposure is associated with abnormal arousal and attention in children. Since methylphenidate (MPD) is widely used to treat attention disorders, we wanted to determine whether prenatal cocaine exposure affects brain function in response to MPD as measured by glucose metabolism in a rodent model. Pregnant rats received 60 mg/kg cocaine or vehicle from gestational days 8-22 by intragastric intubation. On a single day between postnatal days 41-45, offspring received 10mg/kg i.p. of MPD or saline. After 15 min, the quantified 2-deoxyglucose (2DG) method was carried out in freely behaving animals. Seventy nine brain regions were assessed but we focused on functional units such as the mesolimbic and motor circuits which were analyzed using mixed linear models. MPD increased glucose metabolism in most brain regions from 15% to 30% over saline regardless of the prenatal treatment. Prenatal cocaine produced insignificant effects on the rates of brain glucose metabolism overall but produced a reduced response to MPD in the nucleus accumbens in a rostral/caudal gradient compared to control. In addition, correlations of rates of metabolism in the mesolimbic and nigrostriatal systems with the amount of MPD-induced behavior (stereotypy and locomotion) show that prenatal cocaine alters the relationship between regional metabolism and behavior in sex-specific ways. In summary, prenatal cocaine has minimal effects on brain metabolic activity even under drug challenge conditions but has a major impact on the relationship between brain metabolism and behavior.

The multifaceted effects of oral administration of methylphenidate in juvenile rats: anxiety, activity, and attention.

In previous studies, acutely administered oral methylphenidate (MPD, 3mg/kg) prior to testing improved performance on the radial arm maze in juvenile rats. In order to examine the mechanisms producing this improvement we administered MPD once prior to each test of anxiety, locomotor activity and attention. On postnatal day (PND) 22 on an elevated plus maze, rats spent more time beyond the rails on the open arms and showed altered risk-assessment behaviors suggesting an anxiolytic-like effect of MPD. Grid crossings on the plus maze indicated that MPD increased locomotor activity, as did activity recording on PND 23. In another group of juveniles, MPD improved performance in a multi-trial attention task in an age-dependent fashion. These data suggest that oral MPD has multifaceted effects on juvenile rats that together improve performance on cognitive tests such as the radial arm maze. In addition, our data support human studies finding multifaceted effects of MPD.

Sex differences in the effects of cocaine abuse across the life span.

Cocaine alters brain function from the early days of development throughout the entire life of an individual. Since the first preclinical research on cocaine sensitization was published, sex differences in response to the drug in adult rats have been noted. With the appearance of reports on "crack babies" during the 1980s, sex differences in response to prenatal (developmental) exposure have been identified in both clinical and preclinical reports. Cocaine administered during early development in the rat produces wide-spread alterations in function which depend on the timing of drug administration as well as the sex of the animal. In males, the response patterns following postnatal days (PND) 11-20 cocaine administration (equivalent to the late prenatal period in humans) are quite similar to those seen following prenatal exposure (equivalent to the first half of pregnancy in humans). There is a general decrease in dopaminergic (DA) markers and reactivity perhaps due to the uncoupling of the D1 receptor from its second messenger system. While similar changes in D1 uncoupling are seen in females, behavioral and metabolic responses to drug challenges generally show increases in DA responsivity (except adolescents) perhaps due to the activational effects of estrogen and/or decreases in serotonin (5-HT) mediated regulation of DA function. We have found that a significant factor in the hyper-responsivity of the female is the role of the testing environment and the responses to stress which can obscure underlying neurochemical dysregulation. Whether parallel factors are operational in adult males and females is currently under investigation.

Methylphenidate improves performance on the radial arm maze in periadolescent rats.

Methylphenidate (Ritalin; MPD) is one of the most commonly prescribed drugs in childhood and adolescence and many clinical studies have documented its efficacy. Due to the limitations of conducting invasive research in humans, animal models can be beneficial for studying drug effects. However, few animal studies have demonstrated the effects of methylphenidate on cognitive processes. The objective of this study was to find a dose of methylphenidate that was effective in improving performance on a spatial working memory cognitive task when administered orally to periadolescent rats. Therefore, we dosed subjects with methylphenidate at 1 or 3 mg/kg/day via gastric intubation from postnatal day 22 to 59 and assessed the effects of the drug on performance on the radial arm maze each day. To enhance performance overall, a second experiment was conducted where the subjects were moderately food restricted (to 90% of the free-feeding weight). Results of Experiment 1 show that during the first week of testing only the 3 mg/kg MPD-treated males showed improved performance (entries prior to repeated entry) when ad lib fed and housed in pairs while the same dose significantly improved performance in both males and females under conditions of food-restriction and individual housing in Experiment 2. MPD also produced a pattern of increased errors and arms entered during the first week, especially in Experiment 2. MPD increased locomotor activity when tested at postnatal day 60 in both experiments. The data suggest that 3 mg/kg oral methylphenidate improves performance on a spatial cognitive task only early in treatment in the rat. While males show improvement under conditions of both high and low motivation, females only show MPD effects when highly motivated. Hypothetically, methylphenidate may improve radial arm maze performance through increased attention and improved spatial working memory and/or alterations in locomotion, reactivity to novelty or anxiety. Regardless, the study supports the utility of the rat as a suitable model to examine the effects of low dose oral MPD.

Oral methylphenidate improves spatial learning and memory in pre- and periadolescent rats.

Methylphenidate (MPD) is widely prescribed for attention-deficit/hyperactivity disorder in the United States. Patients, mostly school-age children, are taking the drug orally. To simulate the human condition, the authors used a cracker to administer methylphenidate orally (without the stress of handling) from Postnatal Day (PND) 22 to PND 40 and determined the effects of daily low-dose administration on the learning and performance of a radial arm maze win-shift task with all 8 arms baited. Number of entries to repeat, time to finish 8 entries, and days to reach criterion (at least 7 entries without errors for 4 out of 5 consecutive trials) were evaluated. An improvement during the first 7 days was revealed in both male and female rats treated with 3.0 mg/kg of oral methylphenidate compared with the controls. On PND 40, locomotor activity levels were not significantly different in the 3.0 mg/kg treated group compared with the controls during the initial 5 min or during the full 1 hr of recording. These data suggest that oral administration of low-dose MPD improves spatial learning and memory in both male and female preadolescent rats.

Prenatal cocaine dampened behavioral responses to methylphenidate in male and female adolescent rats.

Clinical and animal data point toward deficits in attention and arousal after prenatal cocaine exposure. Since methylphenidate (MPD) is widely used to treat attention disorders, we wanted to determine whether prenatal cocaine (PC) exposure affects the behavioral response to MPD in young rats of both sexes. Pregnant dams received 60 mg/kg of cocaine or vehicle from gestational days 8-22 by intragastric intubations. After delivery, litters were culled to 10 (5 males, 5 females) and fostered. On a single day between PND 41-44 locomotion was recorded in a Plexiglas box within an Accuscan activity monitor after receiving a single injection of 10 mg/kg intraperitoneally of MPD or saline. Rats were also videotaped for analysis of stereotyped behavior. Results showed that MPD administration enhanced locomotion compared to saline injected groups. PC exposure in male rats did not have any effect on the locomotor response to MPD compared to prenatal controls. However, PC-exposed males showed a lower amount of time spent in low intensity stereotypy compared to prenatal control males and both groups of females that received MPD. PC exposure in female rats that received MPD dampened the locomotor response compared to prenatal control females that also received MPD. In conclusion PC exposure dampens the behavioral response to MPD differentially in males and females with an apparent selectivity of locomotion in females and stereotyped behavior in males.

Repeated administration of methylphenidate in young, adolescent, and mature rats affects the response to cocaine later in adulthood.

RATIONALE: Previous studies have shown that the expression of behavioral sensitization to psychostimulants depends on the age and gender of the animal. OBJECTIVE: This study was conducted to determine the pattern of behavioral sensitization to repeated administration of methylphenidate (MPD) at three different developmental ages and to assess the response to a cocaine challenge in adulthood. METHODS: We gave five daily i.p. injections of 10 or 20 mg kg(-1) of MPD (10 MPD, 20 MPD) or saline to male and female rats beginning on postnatal days (PND) 21, 45, or 60. When all groups reached PND 90, rats were challenged with 10 mg kg(-1) cocaine. For both MPD administration and cocaine challenge, locomotion and stereotyped behaviors were assessed for 1 h. RESULTS: The 10 MPD dose produced increased locomotion over the other two treatments at all ages. Rats that received 20 MPD showed a decline in locomotion across days with an increase in the time spent in high intensity stereotypy by day 5. Animals treated with 10 MPD showed diverse behavioral responses with adolescents showing somewhat dampened stereotypy than the other two age groups. In response to cocaine, pretreatment with MPD at all ages enhanced the cocaine response and produced qualitatively different patterns of stereotyped behavior for each gender and pretreatment age group. CONCLUSION: MPD produced clear age-specific sensitization of behavior in rats. Furthermore, exposure to MPD cross-sensitized with cocaine regardless of the age at which MPD exposure occurred with each pretreatment age group showing a unique pattern of responses.