Developmental toxicity of sodium fluoride measured during multiple generations.

Sodium fluoride (NaF) has been used to fluoridate drinking water in the United States since the mid 1940s. Because of the lack of reliable studies on the multigeneration effects of the compound, NaF (0, 25, 100, 175 or 250 ppm in drinking water) was given to rats continuously during three generations. Parental (F0) generation rats were treated for 10 weeks and mated within groups. At gestation day 20, caesarean sections were performed and eight F0 females per group and their litters (F1) were observed for implant status, fetal weight and length, sex and morphological development. The remaining F0 females (29-32 per group) were allowed to litter. F1 offspring (36 of each sex per group) were mated within groups, and caesarean sections were performed at gestation day 20. The F1 females and their litters (F2) were observed for implant status, fetal weight and length, sex and morphological development. In addition, F2 fetuses were evaluated for internal (soft-tissue) and skeletal development. Decreased fluid consumption for F0 and F1 dams at 175 and 250 ppm was attributed to decreased palatability of the solution. No dose-related effects in feed consumption or mean body weight gain were observed in either F0 or F1 females. Numbers of corpora lutea, implants, viable fetuses and fetal morphological development were similar in all groups. No dose-related anomalies in internal organs were observed in F2 fetuses. Ossification of the hyoid bone of F2 fetuses was significantly decreased at 250 ppm. Because of the decreased ossification of the hyoid bone, 250 ppm is considered the effect level.

Multigenerational evaluation of sodium fluoride in rats.

Since the mid 1940s, fluoride has been added to tap water in American communities in an effort to reduce the incidence of dental caries in the population. When the levels of fluoride in drinking water were tested and set, water was the only measurable source of fluoride for most communities. Now, adults and children ingest fluoride with foods and beverages prepared with fluoridated water, and they are exposed to fluoride-containing dental products. As a result, exposure to fluoride is greater than had been anticipated. In the early 1990s, the existing reproductive studies were reviewed in several reports and were considered to be inadequate to determine potential reproductive or developmental hazards. The effects of sodium fluoride ingestion at 0, 25, 100, 175 or 250 ppm in drinking water measured in rats throughout three generations are reported here. Feed and fluid consumption, body weights and clinical signs were recorded at regular intervals. Decreased fluid consumption observed at 175 and 250 ppm was attributed to decreased palatability and did not affect reproduction. No cumulative effects were observed in the three generations. Mating, fertility and survival indices were not affected. Organ-to-body-weight ratios and organ-to-brain weight ratios were not affected. Sodium fluoride up to 250 ppm did not affect reproduction in rats.

Testing the potential of sodium fluoride to affect spermatogenesis in the rat.

The potential of sodium fluoride (NaF) to affect spermatogenesis and endocrine function was assessed in P and F1 generation male rats. Male and female experimental rats received sodium fluoride in their drinking water at one of four concentrations (25, 100, 175, 250 ppm). P generation male and female rats were exposed to sodium fluoride in their drinking water for 10 wk and then males were mated to females within the same treatment groups. Reproductive tissues were collected from P generation male rats after approximately 14 wk of treatment. Pregnant females (P) were exposed to sodium fluoride via their drinking water through gestation and lactation. F1 generation weanling male rats remained within the same treatment groups as their parents. F1 generation male rats were exposed to sodium fluoride in their drinking water for 14 wk, at which time reproductive tissues were collected. Dose-related effects were not observed within the P and F1 treatment groups in testis weights, prostate/seminal vesicle weights, non-reproductive organ weights, testicular spermatid counts, sperm production per gram of testis per day, sperm production per gram of testis, LH, FSH or serum testosterone concentrations. Histological changes were not observed in testicular tissues from either the P or F1 generation. We conclude that prolonged exposure to sodium fluoride in drinking water at the doses administered in this study does not adversely affect spermatogenesis or endocrine function in the P and F1 generation male rats.

Effect of intratesticular injection of sodium fluoride on spermatogenesis.

The potential of sodium fluoride to affect spermatogenesis in the rat was assessed by intratesticular injection. Experimental rats' left testis was injected with sodium fluoride (50, 175 and 250 ppm) in vehicle (0.9% physiological saline); control testes were injected with vehicle. The right testis served as a non-injected control. Testicular tissues collected 'at' and 'distal to' the injection site and from the non-injected control testes were evaluated microscopically 24 hr and 1, 2 and 3 wk post-injection. Testicular tissues obtained at and distal to the injection site in all fluoride-injected groups resembled tissues collected from corresponding areas in the controls. Seminiferous tubule damage observed in both the vehicle-injected control testes and the fluoride-injected testes but not in the non-injected testes was attributed to injection trauma. Polymorphonuclear leucocyte infiltration was observed 24 hr post injection only at the injection site in the vehicle- and fluoride-injected groups. Leydig cells were unaffected. Leucocyte infiltration with seminiferous tubule damage was not considered to be a fluoride treatment-related effect because it was observed in both vehicle- and fluoride-injected testes. The results demonstrate that the rat is not adversely affected by direct exposure to fluoride at levels 200 times greater than those under normal conditions.

Developmental toxicity of sodium fluoride in rats.

Despite the chronic exposure of the US population to fluoridated drinking water since the 1940s, existing studies have been judged inadequate to determine any potential reproductive or developmental hazard. This study was conducted to determine the effects of sodium fluoride (NaF) on foetal development. Sperm-positive female rats were given 0, 10, 25, 100, 175 or 250 ppm NaF daily throughout gestation. They were dosed by drinking water to mimic human exposure to fluoridated water. No dose-related behavioural changes or maternal clinical signs were noted. Fluid consumption by females in the 175- and 250-ppm groups was significantly less than that of the control females. Because of this decreased fluid consumption, the daily amount of NaF ingested (0, 1.4, 3.9, 15.6, 24.7 and 25.1 mg/kg body weight) was less than expected at the two high levels. Feed consumption decreased significantly at 250 ppm, and body weights of pregnant females reflected feed consumption trends. The mean number of viable foetuses per female in all treated groups was similar to that of the control group. The significant decrease in the mean number of implants per litter in the 250-ppm group is probably linked to the lower mean number of corpora lutea in this group. The occurrence of in utero deaths was similar in the control and treated groups. Foetal growth (in terms of foetal body weight and crown-rump length) was not affected by NaF, despite the fact that the dams in the 250-ppm group ate significantly less feed and drank significantly less fluid. There was no dose-related increase in the number of external anomalies in foetuses due to NaF ingestion. At the doses given, NaF had no effect on the development of specific bones, including sternebrae. A significant increase was seen in the average number of foetuses with three or more skeletal variations in the 250-ppm group; the number of litters with foetuses with three or more skeletal variations was increased in the 250-ppm group also, but the increase was not significant. There was no dose-related effect of NaF on the incidence of soft tissue variations.

Mineral interactions in rats fed AIN-76A diets with excess calcium.

The effects of moderate increases in dietary calcium on maternal and foetal mineral interactions were studied in Charles River CD/VAF Plus rats. Female rats were given 0.50, 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 wk before mating, during mating and for 20 days of gestation. Inductively coupled argon plasma-atomic emission spectrometry was used to determine mineral levels in the tissues of non-pregnant rats after 42 days on the diets, in the tissues of pregnant rats on day 20 of gestation and in the whole body of day-20 foetuses. The femurs of the non-pregnant and pregnant rats had a dose-related linear increase in calcium content. In livers of the non-pregnant rats, dose-related linear increases in the phosphorus, zinc and magnesium content were observed, but there was a dose-related decrease in the iron content. There were dose-related linear decreases in the iron and copper contents of the kidneys from the non-pregnant rats. In pregnant rats dose-related linear decreases were observed in the iron content of the liver and in the zinc, iron and magnesium contents of the kidney. The foetuses from rats given a moderate increase in dietary calcium had dose-related decreases in the whole-body contents of phosphorus, iron, copper and magnesium.

Foetal development in rats fed AIN-76A diets supplemented with excess calcium.

This study was designed to evaluate the developmental effects of moderate dietary calcium increases in rats fed nutritionally adequate diets. Female Charles River CD/VAF Plus rats were given 0.50 (control), 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 wk before mating, during mating and for 20 days of gestation. On gestation day 20, the animals were killed and caesarean sections were performed. Both the non-pregnant and pregnant rats in the 0.75, 1.00 and 1.25% groups ate slightly more than did the control group during most of the intervals measured, but not all the increases were statistically significant. There was no consistent pattern of increase or decrease in weight gain. No dose-related changes were found in maternal clinical findings, the average number of implantations, resorptions and viable foetuses, or foetal length or weight. Under the conditions of the study, there were no statistically significant increases as compared with the control group in the litter incidence regarding specific external, visceral or skeletal variations of the foetuses. Dietary calcium was neither foetotoxic nor teratogenic at the concentrations used.