Female reproduction during chronic ethanol consumption in rats

This study was undertaken to ascertain ovarian function under the conditions of ethanol withdrawal and continued ethanol treatment to distinguish between a temporary delay in ovarian activity and a permanent suppression of ovarian function. Immature rats were fed the following diets for 16 weeks: a liquid diet containing 5% ethanol, a liquid diet without ethanol (pair-fed controls), a liquid diet with 5% ethanol for eight weeks followed by laboratory chow and water for eight weeks, or chow and water ad lib. Vaginal patency was significantly delayed in both groups of ethanol-treated rats compared to controls. The duration of an estrous cycle for the rats in the ad lib group was 5.0±0.3 days, while a “regular” estrous cycle was four to six days in duration. The rats which received ethanol for 16 weeks exhibited more irregular estrous cycles (both<4 and >6 days) than the rats with other treatments and the cycles were significantly longer. After 16 weeks of treatment, the rats were mated; ethanol was not given during pregnancy. The average number of pups per litter and body weight of the offspring were similar for all groups. These data show that although ethanol alters normal cyclic activity, it does not totally suppress ovarian function since alcohol-treated rats were capable of mating and delivering viable offspring.     

Chronic ethanol intake modifies estrous cyclicity and alters prolactin and LH levels

The effect on the estrous cycle, as well as prolactin, luteinizing hormone (LH) and alcohol levels, were studied in female rats during chronic alcohol intake. Rats were fed the following diets for 5 weeks: a liquid ethanol diet (5% ethanol, w/v), an isocaloric liquid diet (pair-fed) or laboratory rat-chow and water ad lib. Ethanol-fed rats showed an irregular estrous cycle with a significant decrease in the frequency of the estrus + proestrus phases, and an increase in the duration of the diestrus + metaestrus phases, when compared with both the pair-fed and rat-chow groups. In these alcoholic rats, the estrous cycle phase was correlated with blood alcohol levels, which were found to be lower in the estrus + proestrus than in the other phases of the cycle. At the same time plasma prolactin levels were higher and plasma LH levels lower in the alcoholic rats than in either control group. These data indicate that chronic ethanol consumption prolongs the diestrus phase together with altered plasma prolactin and LH levels in female rats.     

Effect of sodium arsenite on plasma levels of gonadotrophins and ovarian steroidogenesis in mature albino rats: duration-dependent response

Effect of arsenic on ovarian steroidogenesis at the dose available in drinking water at wide areas of West Bengal is reported here. Weights of ovary, uterus and vagina along with biochemical activities of ovarian delta 5-3 beta-hydroxysteroid dehydrogenase (delta 5-3 beta-HSD) and 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) and plasma levels of LH, FSH and estrogen were measured in mature rats of the Wistar strain at diestrous phase following subchronic treatment with sodium arsenite at a dose of 0.4 ppm/rat/day for 16 days (4 estrous cycles) and 28 days (7 estrous cycles). A significant reduction in plasma levels of LH, FSH and estrogen along with significant diminution in the activities of ovarian delta 5-3 beta-HSD and 17 beta-HSD were observed following sodium arsenite treatment for 28 days. This duration of treatment also resulted in a marked degree in diminution in the weights of ovary, uterus and vagina, but 16 days of treatment did not exhibit any significant effect on these above parameters. Arsenic-treated rats exhibited a prolonged diestrous phase in the estrous cycle in contrast to control rats having 4 days of a regular estrous cycle. Deposition of arsenic in ovary, uterus, vagina and plasma was also monitored in arsenic-treated rats. The results of our experiment suggest that duration of arsenic treatment is the critical factor for its adverse effect on ovarian activities at the dose within the range noted in drinking water at several areas of West Bengal in India.     

Flumazenil but not nitrendipine reverses the increased anxiety during ethanol withdrawal in the rat

After 7 day's gradual introduction of ethanol, rats were maintained for a further 4 weeks on a liquid diet containing 10% ethanol (mean daily dose 11.8±0.2 g/kg/day). Control-treated rats received liquid diet alone. Pairs of rats were tested in the social interaction test of anxiety 8 h after withdrawal. Withdrawal from ethanol significantly reduced the time spent in social interaction compared with controls, indicating an anxiogenic withdrawal response. Nitrendipine (50 mg/kg) had no effect on, whereas flumazenil (4 mg/kg) significantly reversed, this withdrawal response. This reversal appeared to be long-lasting as there was still no evidence of increased anxiety when rats were again withdrawn after 3 more days of ethanol diet.     

Similar anxiety-like responses in male and female rats exposed to repeated withdrawals from ethanol

Previous research indicated that male rats exhibited anxiety-like behavior following withdrawal from chronic ethanol exposure. This behavior, as well as other symptoms of withdrawal such as seizure susceptibility, may be sensitized in male rats repeatedly withdrawn from chronic ethanol exposure. Because there are sex differences in some alcohol effects, the present study explored whether male and female rats would respond differently to a course of repeated ethanol withdrawals. Similarly aged male and female rats were exposed to a control liquid diet or a diet containing ethanol (7% w/v). Ethanol-exposed rats had only one 5-day cycle of exposure or three cycles, with 2 days of control diet (CD) between cycles. At 5 h after the final ethanol was removed, the rats were placed as same-sexed pairs in the social interaction test; approximately half of the rats were tested later in the elevated plus maze. Rats withdrawn from ethanol after three cycles exhibited reduced the time spent in social interaction and general activity in the social interaction test and reduced the open and closed arm entries in the elevated plus maze. There were no sex differences in these effects. However, male rats exhibited a small anxiety-like response after one cycle of 5 days' exposure to ethanol and female rats did not. Thus, there are no sex differences in the three-cycle multiple-withdrawal paradigm, but there may be differences after briefer exposures.     

Effect of edifenphos on compensatory ovarian hypertrophy, follicular kinetics and estrous cycle in hemicastrated rats

Edifenphos, an organophosphate fungicide, was administered to hemicastrated albino rats intraperitoneally for 15 consecutive days in graded doses of 2 to 8 mg/kg body weight. In the oil-treated hemicastrated control group, ovarian weight and total number of healthy and atretic follicles were significantly higher than the same parameters in sham-operated control animals, and the estrous cycle was normal. Treatment with 2, 4, 6, or 8 mg/kg/d edifenphos significantly decreased ovarian weight (-6.75, -9.79, -18.71, and -34.13 percent, respectively) below that of the controls. Although treatment with 2 or 4 mg/kg/d edifenphos did not change the number of healthy and atretic follicles, a dosage of 4 mg/kg/d significantly decreased the number and duration of estrous cycles. Treatment with 6 or 8 mg/kg/d, however, significantly decreased the number and duration of different phases of the estrous cycle as well. In all treated groups, the weights of the thymus and uterus were significantly reduced when compared with those of hemicastrated oil-treated control animals.     

Acute effect of ethanol on IgA immunoreactive cells in the intestine-associated immune system

The purpose of this study was to investigate the acute effect of ethanol on mucosa-associated lymphoid tissue at the level of Peyer's patches and the intestinal lamina propria in female rats and to determine whether this action of ethanol is modulated during the estrous cycle. Adult female rats showing proestrus or diestrus day 1 were treated intraperitoneally (ip) with ethanol (4 g/kg). Untreated and saline-injected rats were used as controls. The animals were sacrificed by decapitation 0.5 h after ethanol administration. Immunoglobulin A (IgA) immunoreactive cells were analyzed by indirect immunohistochemistry using mouse anti-rat IgA and a Dako LSAB+ kit. The number of IgA-immunoreactive cells in Peyer's patches was unaltered by ethanol treatment at both phases of the estrous cycle. However, stereological analysis revealed a significant increase in the number of IgA-immunoreactive cells (p < 0.01) in the intestinal lamina propria following acute ethanol administration at proestrus and on diestrus day 1. The results indicate that the intestinal lamina propria, the effector site of the mucosal immune system, can be affected by a single dose of ethanol at both phases of the estrous cycle.     

Estrous cycle-dependent changes in basal and ethanol-induced activity of cortical dopaminergic neurons in the rat.

The influence of the estrous cycle on dopamine levels in the rat medial prefrontal cortex under basal and ethanol-stimulated conditions was evaluated by microdialysis. The basal dopamine concentration in the dialysate varied markedly during the estrous cycle, being highest in estrus and lowest in proestrus. Furthermore, a challenge intraperitoneal administration of ethanol (0.5 g/kg) induced a significant increase in dopaminergic output (+50%) during estrus but had no effect in diestrus or proestrus. Ovariectomy or pretreatment with either finasteride (a 5alpha-reductase inhibitor) or clomiphene (an estrogen receptor antagonist) prevented this ethanol-induced increase in dopamine concentration. The effect of ethanol was restored in ovariectomized rats by pretreatment with estrogen but not by that with progesterone. Our results thus show that the basal levels of dopamine in the prefrontal cortex are dependent on the phase of the estrous cycle. Furthermore, this dependence appears to be attributable to the effects of ovarian steroid hormones and results in a differential sensitivity of the dopaminergic neurons to ethanol. The hormone-induced changes in the activity of these neurons might contribute to the differences in drug sensitivity and mood state apparent among phases of the estrous cycle and between the sexes.     

Effects of Long-term Ethanol Consumption on Rat Peritoneal Macrophages

Abstract: Rats were fed an all liquid diet for 7–8 weeks. One group received 35% of the calories as ethanol while the other group was pair-fed carbohydrates. Peritoneal macrophages prepared from ethanol treated rats had lower phagocytosis via the Fc-receptor and reduced viability in the presence of endotoxin, but their lysosomal enzyme activities measured (β-glucuronidase, cathepsin D, acid phosphatase and acid DNase) were not different from controls.     

Ethanol stimulation after chronic exposure in C57 mice

The effects of chronic exposure to ethanol via an ethanol containing diet on locomotor activity and on the response to depressive and stimulatory doses of ethanol were examined in C57BL/6cr (C57) mice. Mice maintained for approximately 3 weeks on a liquid diet in which 25% of the calories derived from ethanol had blood levels ranging from undetectable to 60 mg% when sampled at 2200 hr. They were less active than nutritionally equivalent controls, were less depressed by a high dose of ethanol (3 g/kg), and were more stimulated by a low dose of ethanol (2 g/kg). The results establish that chronic EtOH exposure via liquid diets attenuate its depressive effect and exacerbate its stimulatory effect on locomotor activity of C57 mice as previously shown for other strains or for rats. The increased activity elevation in chronically exposed mice may reflect an unmasking of excitation upon developing tolerance to EtOH depression.     

Effects of 1-bromopropane, 2-bromopropane, and 1,2-dichloropropane on the estrous cycle and ovulation in F344 rats.

The present study was performed to investigate comparatively the toxic effects of inhalation exposure of 1-bromopropane, 2-bromopropane, and 1,2-dichloropropane on reproductive physiology, particularly on the estrous cycle and spontaneous ovulation in female F344 rats. The rats received inhalation exposure to different halogenated propanes, and were exposed daily for 8 h throughout almost 3 weeks to 0,50,200 and 1000 ppm of 1-bromopropane or 2-bromopropane, or to 0,50,100 and 200 ppm of 1,2-dichloropropane. Throughout the exposure period of 1-bromopropane or 2-bromopropane, the ratio of the number of estrous cycle of 6 days or longer to the number of all cycles in both 1000 ppm groups were about two-fold the ratio in each control group, however, no significant difference was found between the ratios of exposed and control groups. The ratios of such long estrous cycles in groups exposed to 100 or 200 ppm of 1,2-dichloropropane were six- to seven-fold higher than that of the control group. These ratios in exposed rats differed significantly from those of controls. The number of ovulated ova in rats exposed to 1,2-dichloropropane decreased in a dose-dependent manner, and the number of ovulated ova in the 200 ppm group was significantly different from that of control rats. Such significant changes in ovulation were not observed in rats exposed to 1-bromopropane or 2-bromopropane. The absolute and relative weights of the ovaries and uterus in rats exposed to three halogenated propanes were not significantly different from those in each control. Therefore, the present study clarified that: (1) 1,2-dichloropropane prolonged the length of the estrous cycle and inhibited spontaneous ovulation in F344 rats; and (2) the potency of 1,2-dichloropropane to disturb the female reproductive physiology appeared to be greater compared with that of 1-bromopropane and 2-bromopropane under the present conditions of inhalation exposure.     

Are the effects of chronic ethanol administration on erythrocyte membrane mediated by changes in plasma lipids?

The effect of chronic ethanol consumption on plasma and erythrocyte membrane lipids were studied in rats fed a liquid diet containing ethanol for 4 weeks and intubated with the same diet 90 min prior to killing. Control animals underwent the same treatment, except that their liquid diet did not contain ethanol, but an isocaloric amount of carbohydrates. Plasma total cholesterol, free (unesterified) cholesterol and HDL cholesterol were higher in ethanol-fed animals than in controls. Phospholipids were also higher in the plasma of ethanol-fed animals when compared to controls so that the plasma cholesterol/phospholipid ratio (Ch/PL ratio) of the two groups did not differ significantly, regardless of the cholesterol fraction considered. Experimental and control animals did not differ either in the Ch/PL ratio of their erythrocyte membranes. In view of the fact that it has been suggested that the factor determining the direction of the cholesterol exchanges between plasma and erythrocyte membranes is the equilibrium between their respective Ch/PL ratios, these results are interpreted as being compatible with the hypothesis that the effect of chronic ethanol intake on erythrocyte membrane lipids is mediated through changes in plasma lipids.     

The interaction of ethanol and swimming upon cardiac mass and mitochondrial function

Four groups of female Sprague-Dawley rats received a nutritionally adequate liquid diet formulated for rats. Two groups, one ethanol diet and one control diet swam 6 days/wk for 6 weeks and were designated swim ethanol (SWM-E) and swim control (SWM-C) respectively. Their swimming time increased from 15 min/day on the first day to 2 hrs/day during the final week. One sedentary group received an ethanol diet (SED-E) while another sedentary group received a control diet (SED-C). In the ethanol diet 35% of the calories as ethanol isoenergetically replaced dextrin. The group mean body weights were not different at the end of 6 weeks. The left ventricles of both swimming groups showed similar gains in weight, 13% for the ethanol and 15% for the control. Mitochondrial respiration in the ethanol groups showed a significant depression across substrates and across both populations of mitochondria (subsarcolemmal and intermyofibrillar). The swimming-ethanol interaction in the SWM-E group caused an atrophy of the gastrocnemius-plantaris muscle as evidenced by the 13% loss in weight of the muscle. We conclude that chronic ingestion of ethanol will suppress mitochondrial respiration in sedentary and swimming exercised rats, but will not suppress cardiac hypertrophy in the swimming exercised rats. Muscles that are not chronically overloaded by swimming, such as the gastrocnemius-plantaris muscles will undergo atrophy during the swimming protocol of 6 weeks.     

Sex differences in muscarinic receptor binding after chronic ethanol administration in the rat

Male and female rats were administered ethanol (5% v/v) in a liquid diet for 18 weeks. Pair-fed control animals were fed the same diet except that dextrose was substituted isocalorically for ethanol. Normal controls received a commercial laboratory chow for the same duration. Results showed that, in females, chronic ingestion of an ethanol liquid diet significantly increased the number of muscarinic receptor binding sites compared to both control groups. In contrast, for males, there was no significant difference in the mean number of binding sites among the treatment groups. Furthermore, the mean maximum number of binding sites for males and females varied across brain areas. Males had a significantly greater number of receptor binding sites than females in the striatum, while females had a greater number in the cortex. It was suggested that the geuder differences observed in the present study could be mediated by hormonal effects on central muscarinic functioning.     

The effect of gossypol acetic acid on female reproduction

The effects of gossypol acetic acid (GAA) on various aspects of female rat reproduction were evaluated. Forty sexually mature, nulliparous female Holtzman rats received a daily, oral dose of GAA (20 mg GAA/Kg body wt) for 60 days. Nineteen control animals were utilized. Throughout the treatment period, GAA treated rats had significantly more irregular estrous cycles than did controls, and the mean duration of an estrous cycle in the GAA rats was significantly longer than that of controls. A significantly lower body weight gain was observed in all GAA treated animals. A prolonged time for mating, a decrease in incidence of pregnancy, and a reduction in number of viable embryos at Day 13 of gestation were noted among GAA treated animals. Weaning weights of offspring from GAA dams were significantly less than weaning weights of offspring from control dams.     

Ovotoxicity and PPAR-mediated aromatase downregulation in female Sprague-Dawley rats following combined oral exposure to benzo[a]pyrene and di-(2-ethylhexyl) phthalate

The aim of the present study was to determine the ovotoxicity of female Sprague-Dawley (SD) rats exposed to benzo[a]pyrene (B[a]P) and di-(2-ethylhexyl) phthalate (DEHP), either alone or in combination; the molecular mechanism and the combined effects were also evaluated. Female rats were given intragastric administration of control (corn oil), B[a]P (5 and 10mg/kg), DEHP (300 and 600 mg/kg) and B[a]P+DEHP (at 5mg/kg and 300 mg/kg respectively, or at 10mg/kg and 600 mg/kg respectively) on alternate days for 60 days. Relative ovary weight, estrous cycle, 17β-estradiol blood level, ovarian follicle populations, granulosa cell apoptosis, and gene and protein expression of P450Arom and PPAR were investigated. Our study demonstrated that the combination of B[a]P and DEHP exerts ovotoxicity in female rats and suppression of sex hormone secretion and homeostasis, which is associated with prolonged estrous cycles, decreases in ovarian follicle populations and granulosa cell apoptosis involving a PPAR-mediated signaling pathway of action of the two chemicals. In addition, based on qualitative assessment of the combined toxicity, no interaction effects were observed following combined B[a]P and DEHP administration.     

Accumulation of organotins in seafood leads to reproductive tract abnormalities in female rats

Organotins (OTs) are environmental contaminants used as biocides in antifouling paints that have been shown to be endocrine disrupters. However, studies evaluating the effects of OTs accumulated in seafood (LNI) on reproductive health are particularly sparse. This study demonstrates that LNI leads to impairment in the reproductive tract of female rats, as the estrous cycle development, as well as for ovary and uterus morphology. Rats were treated with LNI, and their reproductive morphophysiology was assessed. Morphophysiological abnormalities, such as irregular estrous cycles, abnormal ovarian follicular development and ovarian collagen deposition, were observed in LNI rats. An increase in luminal epithelia and ERα expression was observed in the LNI uteri. Together, these data provide in vivo evidence that LNI are toxic for reproductive morphophysiology, which may be associated with risks to reproductive function.     

Effects of ethanol and pantothenic acid on brain acetylcholine synthesis.

1. Measurements of brain acetylcholine (ACh) synthesis from precursor [14C]-pyruvate, pantothenic acid (PA) concentration in the brain, and blood ethanol (EtOH) concentration were made in rats treated with either ethanol (5-6 g kg-1 body wt daily) alone or ethanol with PA supplementation (100-200 mg kg-1 body wt daily). EtOH with or without PA was administered orally by either Lieber-Decarli liquid diet for 4 weeks and 4 months or by oral intubation for 1 and 4 days. Matched controls were given either ethanol-free liquid diet or saline. 2. ACh synthesis in the brain of rats treated with ethanol alone for 4 months was significantly (P less than 0.01) inhibited. PA concentration of the brain was diminished to 7.0% of the control value. 3. PA concentration in the brain of rats treated with ethanol plus PA for 4 months was three times that of rats treated with ethanol alone. ACh synthesis in rats with ethanol and PA supplementation was also significantly (P less than 0.01) higher. 4. There was no difference in blood EtOH concentration between rats treated with ethanol with or without PA supplement. 5. The EtOH effect on ACh synthesis and PA concentration in the brain was observed in the chronic treatments but not in the acute treatments. 6. Data suggest that chronic ethanol exposure may decrease ACh synthesis by depleting PA, a precursor for the synthesis of acetyl CoA. Acetyl CoA is an essential substrate for ACh synthesis.     

Transferrin microheterogeneity in rats treated chronically with ethanol

The microheterogeneity of rat serum transferrin was analyzed by isoelectric focusing and immunofixation from Wistar rats chronically treated with ethanol either by inhalation (4 weeks) or by a liquid diet (6 weeks) and from pair-fed controls. In spite of the length of ethanol exposure, a daily ethanol intake of 13-18 g/kg b.wt. and BAC levels of 1.0-4.1 g/l, no transferrin abnormality similar to that found in human alcoholics was observed. It is apparent that these two common animal models for chronic high ethanol consumption are not suitable for experimental studies of ethanol-associated effects on transferrin.     

The Effect of Gossypol Acetic Acid on Female Reproduction

ABSTRACT

The effects of gossypol acetic acid (GAA) on various aspects of female rat reproduction were evaluated. Forty sexually mature, nulliparous female Holtzman rats received a daily, oral dose of GAA (20 mg GAA/Kg body wt) for 60 days. Nineteen control animals were utilized. Throughout the treatment period, GAA treated rats had significantly more irregular estrous cycles than did controls, and the mean duration of an estrous cycle in the GAA rats was significantly longer than that of controls. A significantly lower body weight gain was observed in all GAA treated animals. A prolonged time for mating, a decrease in incidence of pregnancy, and a reduction in number of viable embryos at Day 13 of gestation were noted among GAA treated animals. Weaning weights of offspring from GAA dams were significantly less than weaning weights of offspring from control dams.