With increasing clinical use of L-alpha-acetylmethadol.HCl (LAAM), findings of a carcinogenic bioassay could be useful in risk assessment. Initial studies provided a sex-related oral LD50 in B6C3F1 mice, 126 mg/kg for males and 71 mg/kg for females, and changes after treatment with mean doses of 8, 18, and 33 mg/kg for 90 days which included hyperactivity and unchanged growth rate, food intake, and morphology. Twenty-four-month oral doses were 7.6 and 30.1 mg/kg for mice, 3.1 and 9.7 mg/kg for male rats, and 5.7 and 16.6 mg/kg for female rats. LAAM was lethal in high-dose male mice (40% survival) whereas survival rates of 74-78% were similar in other treated and control groups. An 80-90% survival rate was seen for rats. Deaths related to aging and concomitant morphologic changes occurred randomly in all groups. The causes of LAAM-induced deaths were not established. Central nervous system stimulation and fighting were more common to male rodents. A dose-related inhibition of growth was also common but food intake was stimulated in male mice and both sexes of rats. An increase in liver neoplastic nodules was dose related and drug related in rats. Some nonneoplastic lesions may have been drug related. Terminal plasma LAAM and metabolite levels were generally dose related.

The extensive clinical use of methadone encouraged the performance of a carcinogenesis bioassay to support risk assessment in man. An oral LD50 of 178 mg/kg was obtained in B6C3F1 mice. Physiologic changes induced by mean oral doses of 15, 30, and 60 mg/kg for 90 days included dose-related central nervous system (CNS) stimulation, fighting, tolerance development, sex-related alteration of food consumption, and no drug-related pathology. In the chronic study dosages were 15 and 60 mg/kg for mice, 16 and 28 mg/kg for male rats, and 46 and 88 mg/kg for female rats. Survival incidences for treated and control rodents were 72-86% for mice and 80-90% for rats. Deaths related to morphologic changes of aging occurred in all groups. CNS stimulation and fighting were more common to male rodents. Growth rates were unchanged for mice but a dose-related inhibition occurred for rats. Higher doses stimulated food intake in both species. Neither the type nor incidence of neoplasia was drug related but a few nonneoplastic lesions may have been. Preliminary plasma methadone levels at necropsy were dose related in the rat.

The rationale employed in the design of new chelating ligands for strong binding of Fe(III) at physiological pH for the treatment of iron overload disease such as Cooley's anemia is explained and the preparation and evaluation of forty new iron(III) chelators are described. The new ligands investigated include the following classifications: five analogs of ethylenediaminetetraacetic acid (EDTA) with amino and carboxylate donor groups; ten ligands with phenolic (or potential phenolic) donor groups in addition to amino and carboxylate donors: five ligands containing phenolic groups substituted on pyridine rings in addition to amino and carboxylate donors; six aminophosphonic acid or ester groups with additional phenolate and amino donors; eight macrocyclic polyamines containing auxiliary carboxylate and/or phenolate donor groups; three trishydroxamic acids; two triscatechols; and one multi- dentate ligand with coordinating amide groups. These chelating agents were administered to male BDF1 hybrid mice overloaded with iron by blood transfusion. Fecal and urinary elimination of iron were measured, as well as the retention of iron in the spleen and liver. The results are compared with the action of desferriferrioxamine B (DFB). Potencies relative to a Desferal® dose of 250 mg/kg tested simultaneously and assigned a potency of 1.0 are reported, and relative toxicities (LD50s and toxic signs) are also described. Nine of the ligands tested showed sufficient potency to warrant further development as iron chelating drugs, while six of them have potencies comparable to or greater than that of DFB.

Because of a projected pilot study with EDHPA in Cooley's anemia patients, animal studies with emphasis on reversibility of potential toxic signs were performed. Young dogs were treated iv with 6-18 mg/kg or orally with 30-240 mg/kg for 14 days followed by a 16-day recovery period. Drug-induced emesis, elevated BUN changes in kidney, spleen, and thymus weights diminished during recovery. One deceased dog exhibited nephrotoxicity consisting of tubular necrosis and deposition of the iron-EDHPA complex. The latter was observed in the excreta of survivors but kidney damage was not evident. Atrophy of the spleen and thymus in the deceased dog was consistent with less intense organ weight changes in recovered survivors. In the absence of morphologic changes after recovery, the precise effect on the immune system is unknown. The iv LD50 was 53 mg/kg for rats and mice. No rodent deaths occurred at an oral dose of 6000 mg/kg. An elevated BUN and changes in kidney, spleen, and thymus weights were confirmed in rodents given iv doses of 5-20 mg/kg or oral doses of 150-600 mg/kg for 5 days. It is cautioned that during the use of EDHPA derivatives that the functions of the renal and immune systems be monitored.

Few teratogenic studies in animals have been performed simulating marihuana smoking in man. An inhalation marihuana teratology study was conducted in albino rabbits utilizing a modified automatic smoking machine originally developed for rats and mice. Appropriate numbers of dams were exposed to 4 puffs (0.14 mg/kg), 8 puffs (0.72 mg/kg), or 16 puffs (1.44 mg/kg) once daily during gestation Days 6 to 18, and sacrificed on Day 28. Control dams were exposed to 12 puffs of placebo cigarettes or sham-treated for a similar duration in the absence of any smoke. Consistency of smoke was monitored by cigarette weights, total particulate matter, concentrations of carbon monoxide (CO), and tetrahydrocannibinol (THC) in smoke, carboxyhemoglobin levels, and plasma THC levels. Except for a transient decrease in dam respiration rates, other gross toxic signs were absent. Reproductive parameters of mothers were generally normal except for a dose-related embryotoxicity predominantly associated with early resorptions. Despite twice the number of embryo/fetal deaths, there were no marihuana soft tissue or skeletal defects. A correlation between dam demises and CO levels among placebo-exposed animals was related to greater quantities of CO being generated during placebo combustion. It has been shown in the rabbit that marihuana is embryotoxic and not a teratogen at plasma THC levels found in human females.

Because of potential use as therapeutic agents, additional toxicologic studies on pure cannabinoids and cannabis extracts were conducted. Monkeys received a single iv dose of cannabidiol (CBD) or cannabichromene (CBCH) as an aqueous emulsion and LD50s were 212 and 270 mg/kg, respectively. The larger doses of CBD elicited tremors, convulsions, hypopnea, bradycardia, and cardiac failure. Survivors from smaller doses recovered in 1–3 days and liver weights increased 19–142%. CBCH evoked a dose-related hyperpnea and salivation initially but subsequent CNS inhibition resulted in apnea and bradycardia. Liver weights were elevated 8–71% and kidney weights 10–155% and pulmonary irritation, nephritis, increased BUN, and decreased serum electrolytes, hemoglobin, and RBC were found. Hashish oil containing 11.6 or 31.1% Δ9-THC caused muscle spasms, salivation, dyspnea, arrhythmia, and hypothermia and the LD50s were 326 and 435 mg/kg, respectively. Survivors recovered in a few days; lung and spleen congestion were common. Five day iv treatment with hashish oil 65–260 mg/kg) initiated similar toxicity and tolerance developed by Day 4. Growth rates were inhibited 7–20% and liver, kidney, and heart weights increased 20–100%. Delayed lethality occurred in two monkeys; histopathology included injection site necrosis, hemorrhagic foci in lungs, heart, intestines, and endocrines. Ninetyday oral treatment with CBD (30–300 mg/kg) had little effect but liver and kidney weights rose 13–56% above controls without morphologic changes. Decreased testicular size and inhibition of spermatogenesis occurred. The temporal pattern of adverse responses differentiated CBD, CBCH, and hashish oil.

Responsible evaluation of energy production effects on human health requires prior accounting for the socioeconomic, cultural, and climatic characteristics known to influence mortality rate and cause. Fifteen population characteristics and environmental variables (education, income, occupation, industrial mix, socioeconomic status, housing quality, climate, urban residence, geographic residence, internal migration, cigarette consumption, alcohol consumption, marital status, foreign birth or stock, and religious affiliation) and three age subgroups are discussed. An initial set of eight variables is indicated for mortality rate standardization, based on the reliability of their relationships with mortality. These eight variables are: education, occupation, industrial mix, urban residence, marital status, ethnic mix, and cigarette and alcohol consumption. Education and occupation are negatively related to mortality. Occupational exposure to toxicants (indicated by industrial mix), cigarette consumption, and alcohol consumption have positive linear relationships with various specific causes of mortality. Urban residence, marital status, and ethnicity have non-linear relationships with mortality and show consistent patterns for certain causes of death. In addition to these characteristics three age subgroups ( less than 1 year, 1-14 years, greater than or equal to 65 years) are discussed because of their relatively high or low rates compared to the rest of the population. A brief review of water and air pollution effects on mortality is included for completeness. Unique to this review is the quantitative summary (presented as an appendix) of the variables influencing adult mortality. It is a compilation of numerical relationships, derived either directly or indirectly from the published data, that support the choice of influencing variables.

Exposure of rats to marihuana or placebo smoke for periods up to 365 days was performed with an automatic inhalator. Δ9-Tetrahydrocannabinol (Δ9-THC) concentrations in the marihuana smoke were similar to those inhaled by man and were presented to the rats in a 50-ml puff volume of 2-sec duration and a 30-sec exposure interval followed by a 30-sec period of fresh air each minute (1 puff/min). By varying the number of puffs from three simultaneously smoked marihuana cigarettes (0.9 to 1.2% Δ9-THC), 8 to 10 Fischer rats simultaneously received a single daily Δ9-THC dose of 0.4, 0.8 or 1.5 mg/kg, 6–7 days per week for 365 days. All treatment groups contained 30 males and 30 females except for the high-dose group which had 50 males and 50 females. At each of the lower doses 260 (3%) of the animals died while at the higher dose 18108 (17%) died. Twenty-five percent (1560) of the placebo-smoked rats died primarily from carbon monoxide poisoning. No sham-smoked rats died. In deceased marihuana-smoked rats, organ congestion and focal petechial hemorrhages in the brain suggested circulatory failure. In contrast with an earlier 87-day inhalation study, pulmonary irritation progressed beyond a dose-related focal alveolitis or pneumonitis with the accumulation of yellow-brown alveolar macrophages admixed with a few neutrophils and mononuclear cells, to a spectrum of more pronounced inflammatory and focal proliferative changes. The development of focal granulomatous inflammation in the lung with giant cell forms of macrophages and cholesterol-like clefts were striking new developments in the marihuana-smoked rats, especially since these effects were dose related and nonreversible after a 30-day recovery period.

Groups of male and female Fischer rats were exposed to marijuana cigarette smoke via an automatic smoking machine. Inhaled Δ9-tetrahydrocannabinol doses of 0.7, 2, and 4 mg/kg were relevant to man and were given for 12, 18, 27, 57, and 87 days. Another group of rats treated for 87 days was studied after a recovery of 20 days. Control animals inhaled smoke produced by placebo cigarettes. In the first week of exposure, 20% of lower-dosed rats were hyperactive and 50% at the high dose were prostrate or ataxic upon removal from the inhalator. Behavioral aberrations ameliorated within a few hours except for the depression exhibited by males at the high dose. Tolerance to CNS inhibition developed in 1–2 weeks. CNS stimulation, as manifested by hypersensitivity and hyperactivity, progressively involved more animals, primarily females, in all groups during Days 27–57. Tolerance to CNS stimulation developed thereafter. Fighting was displayed by 90% of females and 50% of males at 4 mg/kg by weeks 6–7. Neurotoxicity was expressed by involuntary vertical jumping, predominantly among high-dosed males in weeks 3 and 8. Normal behavior was displayed after cessation of treatment. At necropsy, homogenates of cerebrum and cerebellum were prepared and were analyzed for protein, RNA and acetylcholinesterase (AChE) activity. Cerebral AChE activity in females increased 33–71% after 12 exposures, decreased 10–23% after 57 exposures and rose 12% after 87 exposures. Cerebellar enzyme activity initially increased 15–35% in animals of both sexes during the subchronic phase but declined in females after 27 exposures. The extent of change in enzyme activity was generally reduced with continued treatment. Cerebellar RNA increased approximately 20% in rats of both sexes, but at different time intervals during the subchronic phase, and remained elevated in females at 87 days. Neurochemical changes were sex related and coincided with behavioral manifestations, and some changes extended into the recovery period. Inhalation findings were similar to those obtained earlier by the oral route; however, females demonstrated a greater facility to adapt to the cumulative toxic effects of marijuana smoke.

Groups of male and female Fischer rats were treated orally with Δ 9 -THC doses of 2, 10, or 50 mg/kg for 28, 90, and 180 days. Another group of rats treated for 180 days was studied after a 30-day recovery period. The lowest dose employed in the present study corresponded to Δ 9 -THC content of one high-grade marihuana cigarette and the intermediate dose simulated the Δ 9 -THC content of one hashish cigarette. In the first 10 days CNS-depression, incoordination, ataxia and passivity, poikilothermia, and hypopnea occurred to which tolerance developed. During days 10–20, irritability, hypersensitivity, hyperactivity, and aggression predominated. Peaks of fighting occurred between days 20–100 among 60% of the animals at higher doses. Tremors and clonic convulsions appeared after day 70 in 50% of the animals at 50 mg/kg and 12% at 10 mg/kg. Tolerance developed to CNS-stimulation, fighting, and neurotoxicity. At necropsy, different portions of the brain were used to prepare 10% ( w v ) homogenates which were analyzed for protein, RNA, and AChE activity. After 28 treatments, only cerebellar RNA increased approximately 16% in both sexes on the high dose. RNA was unchanged after 90 treatments although there was a parallel decline in RNA and protein in various parts of female brain. An 11–21% decrease in RNA was found in cerebral portions of males receiving 50 mg/kg for 180 days. A dose-related fall in protein and RNA in cerebral and cerebellar homogenates from both sexes resulted in no alteration of RNA concentration after a 30-day recovery interval. AChE activity was depressed 17–34% in female cerebellum after 28 treatments and continued treatment for 90 days initiated slight decrements in enzyme activity in various portions of male brain at the high dose. Prolonged treatment for 180 days elevated AChE activity 19–70% in all portions of brain in the males in a nearly dose-related fashion. On the other hand, AChE activity diminished 8–48% in the female. During the recovery period, enzyme activity increased 11–43% in cerebrum and 19–78% in cerebellum for both sexes. Neurochemical alterations were in part sex-related and coincided with behavioral changes associated with cumulative toxicity. Although tolerance is known to develop to most clinical abnormalities, some neurochemical changes persisted after cessation of treatment.

The chronic toxicity of Δ9-tetrahydrocannabinol (Δ9-THC) given orally 28, 90, and 180 days to Fischer rats at doses of 2, 10, and 50 mg/kg was investigated. Some 180-day treated animals were monitored after a 30-day recovery interval. The lower doses used corresponded to the Δ9-THC content of marihuana or hashish. In the first 10 days CNS-depression, incoordination, ataxia and passivity, poikilothermia, and hypopnea occurred to which tolerance developed. During days 10–20, irritability, hypersensitivity, hyperactivity, and aggression predominated. Fighting occurred between days 20–100. Tremors and clonic convulsions appeared after day 70 in 50% of the animals at 50 mg/kg and 12% at 10 mg/kg. Tolerance developed to CNS-stimulation, fighting and neurotoxicity and lethal cumulative toxicity was seen although the cause of death was not established. Growth rate of both sexes was inhibited despite an elevation in food consumption after a transient anorexia No morphological changes could be ascribed to Δ9-THC. Except for a rise of 28–45% in SGOT and 46–69% in SGPT in males at higher doses and a belated hyperglycemia in both sexes, clinical chemistry, hematological, and urinalysis parameters were within normal ranges. The greatest changes were seen in ratios of organ/FBW at 50 mg/kg: 10–20% increase in brain, lungs, kidneys, heart, and liver; 45% increase in adrenals; 96% increase in male pancreas and 13–25% increase in testis and prostate. The present investigation implicated reasonable doses of Δ9-THC in undesirable behavioral changes highlighted by fighting aggression, convulsive activity, and lethal cumulative toxicity. The absence of morphological changes despite changes in growth rate and organ weights indicated a functional impairment that was not an immediate threat to the life of the organism because of initiation of as yet unknown protective mechanisms.

Subcutaneous (s.c.) administration of delta-9-tetrahydrocannabinol (delta-9-THC) to rabbits produced dose-related cumulative toxicity. Five groups of three New Zealand albino rabbits each received 28 daily treatments with isotonic saline, sesame oil of 15.9, 45.0 or 153.4 mg/kg/day of delta-9-THC dissolved in sesame oil. Dose-related dermal responses included erythema, edema, ulceration and nodule formation. Some of the granulomatous nodules contained an oily substance and exhibited liquefactive necrosis. The severities of erythema and ulceration were generally maximal during the first week of treatment, but edema and nodule formation were most severe after days 12 and 14, respectively. All rabbits survived treatment, but body weights, liver weights and liver glycogen levels were decreased in a dose-related manner. Maximal body weight effects occurred after day 19. Hemochemical changes occurred only in rabbits treated with 153.4 mg/kg/day and included decreased blood sugar and alkaline phosphatase, and increased serum potassium. Hematology parameters were normal throughout the treatment period. No drug-related pathological lesions occurred in internal organs. The cumulative body weight changes, significantly decreased hepatic glycogen levels and reduced blood sugar and alkaline phosphatase values may have indicated drug-induced metabolic changes.

Exposure of rats to marihuana or placebo smoke for periods of up to 87 days was performed with an automatic inhalator. Δ9-Tetrahydrocannabinol (Δ9-THC) concentrations in the marihuana smoke were similar to those inhaled by man and were presented to rats in a 50-ml puff volume of 2-sec duration and a 30-sec exposure interval followed by a 30-sec period of fresh air each minute. By varying the number of puffs from three simultaneously smoked marihuana cigarettes (2.1% Δ9-THC), 8–10 Fischer rats simultaneously received a single daily Δ9-THC dose of 0.7, 2, or 4 mg/kg, 6 consecutive days per week for 27, 57, or 87 days. Lethal cumulative toxicity in male rats began in the first week, which eventually resulted in 60% deaths, at two peak intervals, in weeks 3 and 8. During the first week, some dose-related CNS inhibition, hypothermia, and hypopnea occurred to which tolerance developed at different rates. In the second and third weeks, CNS stimulation was prevalent at lower doses. Neurotoxicity (“popcorn” reaction) occurred in 70% of both sexes on the high dose and reached a peak in weeks 3 and 8. Tolerance to these manifestations developed in subsequent weeks. Generally, growth rates and food intake decreased in males treated with the high dose but, in the other treated groups, food and water consumption were evaluated throughout the study. Increased organ to body weight ratios for the brain, lungs, and heart of males reflected decreases in final body weight. No convulsive episodes were noted. In weight-decreased rats, congestion in major organs and pulmonary edema suggested circulatory failure. In the treated animals, which were sacrificed, a dose-related moderate focal pneumonitis, characterized by the accumulation of aggregates of yellow-brown, sudanophilic alveolar macrophages, polymorphonuclear leukocytes, and lymphocytes, was observed. Female rats evoked as yet unknown protective mechanisms more efficiently than males in the face of neurotoxicity and morphological changes.