Animal evidence considered in determination of cannabis smoke and Δ9 -tetrahydrocannabinol as causing reproductive toxicity (developmental endpoint); Part I. Somatic development.

OBJECTIVES: On December 11, 2019, California's Developmental and Reproductive Toxicant Identification Committee (DARTIC) met to consider the addition of cannabis smoke and Δ9 -THC to the Proposition 65 list as causing reproductive toxicity (developmental endpoint). As the lead state agency for implementing Proposition 65, the Office of Environmental Health Hazard Assessment (OEHHA) reviewed and summarized the relevant scientific literature in the form of a hazard identification document (HID). Here we provide reviews based on the HID: shortened, revised, and reformatted for a larger audience. METHODS: While the HID included both human and animal data, this set of three reviews will highlight the animal-derived data pertaining to somatic development (Part I), neurodevelopmental effects (Part II), and proposed neurodevelopmental mechanisms of action (Part III). RESULTS: Endogenous cannabinoids (eCBs) and their receptors serve many critical functions in normal development. Δ9 -THC can interfere with these functions. Mechanistic studies employed techniques including: blocking Δ9 -THC binding to endocannabinoid (EC) receptors, inhibiting Δ9 -THC metabolism, and/or using animals expressing knockout mutations of EC receptors. Apical somatic effects of cannabis smoke or Δ9 -THC reported in whole animal studies included decreases in offspring viability and growth. Mechanistic studies discussed in Part I focused on Δ9 -THC effects on early embryos and implantation, immune development, and bone growth. CONCLUSIONS: In reaching its decision to list cannabis and Δ9 -THC as a developmental toxicant under California's Proposition 65, the DARTIC considered biological plausibility and the consistency of mechanistic information with effects reported in human and whole animal studies.

Animal evidence considered in determination of cannabis smoke and Δ9 -tetrahydrocannabinol (Δ9 -THC) as causing reproductive toxicity (developmental endpoint); Part II. Neurodevelopmental effects.

This review focuses on neurodevelopmental effects observed in animal studies of cannabis smoke and Δ9 -THC. Effects in offspring after preconceptional, prenatal, or perinatal exposure to cannabis smoke or Δ9 -THC were considered. Locomotor and exploratory behavior effects were noted in rats. Cognitive effects observed included impairment of memory and learning, attention deficits, time taken to complete tasks (rats) and alterations in response to visual stimuli (rats/monkeys). Emotionality was observed in rodents as an increase in separation-induced ultrasonic vocalizations, reduced social interaction and play behavior, and increased generalized anxiety. Increased rate of acquisition of morphine self-administration and/or enhanced sensitivity towards the rewarding effects of morphine or heroin were observed in adult rats prenatally exposed to Δ9 -THC. Expression of cannabinoid receptors was examined in rodent studies along with behavioral parameters. Altered mRNA levels of genes relevant to synaptic plasticity in the nucleus accumbens (the brain region associated with compulsivity, addiction vulnerability, and reward sensitivity) were noted. Findings in zebrafish supported effects in mammalian models. Neurochemical effects on specific brain regions and neurotransmitter systems seen in these animal studies appear to impact cognitive function, motor activity, and drug sensitivity. Mechanistic studies provided evidence for the biological plausibility of effects observed. Observations from animal studies of changes in motor behavior, cognitive performance, emotionality and susceptibility to drug sensitivity later in life were among the findings from animal and human studies considered by California's Developmental and Reproductive Toxicant Identification Committee, in concluding that cannabis smoke and Δ9 -THC are developmental toxicants.

Animal evidence considered in determination of cannabis smoke and Δ9 -tetrahydrocannabinol as causing reproductive toxicity (developmental endpoint): Part III. Proposed neurodevelopmental mechanisms of action.

This review summarizes the most common potential pathways of neurodevelopmental toxicity due to perinatal exposure to Δ9 -tetrahydrocannabinol (Δ9 -THC) that lead to behavioral and other adverse outcomes (AOs). This is Part III in a set of reviews highlighting the animal-derived data considered by California's Developmental and Reproductive Toxicant Identification Committee (DARTIC) in 2019. The Hazard Identification Document (HID) provided to the DARTIC included a summary of human, whole animal, and mechanistic data on the neurodevelopmental toxicity of cannabis smoke and Δ9 -THC. The literature search for mechanistic data has been updated through 2020. We focus on mechanistic pathways relating to behavioral and other neurodevelopmental outcomes of perinatal exposure to Δ9 -THC. The endocannabinoid system (EC system) plays a crucial role in many processes involved in neurodevelopment and exposure to Δ9 -THC can alter these processes. Whole animal studies report changes in cognitive ability, behavior, and motor function after prenatal exposure to Δ9 -THC. Findings from mechanistic studies add to this evidence and further provide information regarding the pathways leading to these outcomes. Neuromechanistic studies can bridge the gaps between molecular initiating events and apical neurodevelopmental endpoints caused by a chemical. They offer insight into potential alterations in the same pathways by other chemicals that can also result in AOs. Studies of cannabinoid receptor agonist-induced molecular alterations and provide deep biological plausibility at the mechanistic level for the cognitive, behavioral, and motor impairments observed in animal studies after perinatal exposure to Δ9 -THC.

Toxicity endpoint selections for a simazine risk assessment.

BACKGROUND: California uses simazine at one of the highest levels for states in the United States (approximately 2.5 million lbs 2006-2010). Simazine causes neuroendocrine disruption and mammary cancer in test animals. A risk assessment was prioritized by the California Department of Pesticide Regulation because of the nondietary concern for simazine exposure to occupational/nonoccupational simazine users, resident nonusers, and bystanders (especially children and children exhibiting pica) at greatest risk. METHODS: No observed effect levels (NOELs) from animal studies as well as human exposure data were used to determine nondietary values for the above populations. Registrant-submitted and open literature studies focusing on oral (major human route) effects for simazine and the major metabolites desisopropyl-s-atrazine and diaminochlorotriazine were reviewed as part of the hazard identification process. RESULTS: Developmental, reproduction, and chronic studies provided the lowest NOELs for the acute (5 mg/kg/day), subchronic (0.56 mg/kg/day), and chronic (0.52 mg/kg/day) exposure durations, respectively. A benchmark dose (95th percentile) was calculated for mammary tumorigenesis, assuming a threshold mechanism in rats (benchmark dose lower limit [95th percentile; BMDL05 ]: 2.9 mg/kg/day). Margins of exposure and uncertainty factors (100-300×, depending on exposure scenario) were used to characterize risk for designated population subgroups. CONCLUSIONS: Fetal developmental delays, endocrine disruption, and mammary tumors resulted from simazine treatment. Systemic and maternal/fetal effects determined the critical NOELs used in risk assessment. Margins of exposures for most scenarios were below acceptable levels, especially for children who may be bystanders where simazine is applied and children who exhibit pica. This risk characterization raises a concern for long-term effects in humans.

Reduced water intake: Implications for rodent developmental and reproductive toxicity studies.

In developmental and reproductive toxicity studies, drinking water is a common means of delivering the test agent. Reduced consumption of toxicant-containing water raises questions about indirect effects of reduced maternal fluid consumption resulting from unpalatability, versus direct effects of the test compound. Issues to consider include: objective assessment of dehydration and thirst, the relative contributions of innate and learned behaviors to drinking behavior and flavor preference, and the objective assessment of physiologic stress. Not only do lab animals under ad lib conditions consume more water than the minimum required to maintain fluid balance, animals faced with water restriction have substantial physiologic capacity for protection of metabolic processes. Measures of blood biochemistry can provide quantifiable, objective indications of fluid balance, but changes in these parameters could result from other causes such as effects of a test toxicant. Consummatory behaviors in response to perceived need are highly influenced by learning. Hence, the drinking behavior, water intake, and flavor acceptance/preference of animals used in toxicology experiments could be subject to learning experiences with the test compound. Physiological symptoms of stress produced by water deprivation may be distinguishable from the symptoms associated with other generalized stressors, such as food deprivation, but doing so may be beyond the scope of most developmental or reproductive toxicity studies. Use of concurrent controls, paired to test groups for water consumption, could help distinguish between the direct effects of a test toxicant as opposed to effects of reduced water consumption alone.

A dietary risk assessment of the pyrethroid insecticide resmethrin associated with its use for West Nile Virus mosquito vector control in California.

An outbreak of human illnesses associated with West Nile Virus (WNV) occurred in New York City in 1999. Since then, it has gradually spread westwards, reaching northern California for the first time in 2005. WNV is transmitted by several mosquito species and birds serve as the main reservoir. Several control measures have been used, targeting both the aquatic larvae and the adult mosquitoes. In the latter case, roosting birds in trees are sprayed with pyrethroid insecticides because these are highly toxic to mosquitoes, but have low avian toxicity. A request was made to use a resmethrin-containing insecticide during the month of October 2005 in California. Because resmethrin was not registered for use on growing crops, concerns were raised about potential crop contamination. Therefore, an expedited dietary risk assessment was conducted on resmethrin. Developmental toxicity in the rat (NOELs of 25 or 40 mg/kg/day) was used as the acute endpoint and dietary exposure was assessed using the DEEM-FCID computer program. Only crops growing above ground during October were considered. Margins of Safety (MOS) were found to be above 100, the level generally considered to be sufficient to protect public health when using an animal NOEL.

Effects of restraint stress in gestation: implications for rodent developmental toxicology studies.

Restraint has been used as a procedure to study the effects of stress on gestation outcome in rodents. The effects of restraint could potentially be used as a model for the impact of general stress produced by high doses of toxicants and other interventions. In mice, restraint in the peri-implantation period leads to implantation failure, and restraint at appropriate times in organogenesis produces cleft palate, supernumerary ribs, and resorption. In rats, there is some evidence for an association with restraint for implantation failure, but not for the morphological anomalies. Restraint in late gestation alters adult sexual behavior of male rat offspring, but consequences for their fertility are not known. Intrauterine growth retardation is not commonly associated with gestational restraint. In the few studies where they have been directly compared, different restraint procedures produced graded, qualitatively different, or no effects. Adrenocortical hormones have been implicated as mediating the effect of restraint on cleft palate, but not on supernumerary ribs, implantation failure, or sexual differentiation. Given the variety of restraint procedures and the varying species-dependent consequences, it is not possible to infer a generalizable pattern of developmental effects due to gestational stress from the restraint literature. As an alternative approach, contemporary methods in gene expression and developmental biology could profitably be applied to understanding different patterns of stress-mediated effects of toxicant exposures on intrauterine development.