Development of a nurse-provided health system strategy for diabetic foot care.

A nurse-provided university health system diabetic foot screening/education/treatment program evaluated 403 patients in the initial 12 months of development. All patients were provided individualized foot-specific patient education, varying in intensity with the magnitude of their risk status. One hundred and forty-five (36%) were categorized as being at risk for the development of a diabetic foot ulcer. Improper footwear capable of producing foot ulceration was recorded in 268 (66.5%) of the enrollees. Seven patients with previously undiagnosed Charcot foot disorder were identified. Eighty-three of the enrollees were seen at least once in follow-up. Sixty-one (73%) used improper footwear at the initial evaluation, which was decreased to 36 (43%) at the first follow-up visit. Nurse-provided foot-specific diabetic screening and education, combined with protective footwear, has been shown to be a cost- and resource-effective method of decreasing the rate of diabetic foot ulcers, and the risk for eventual lower extremity amputation.

Effect of chronic ethanol exposure on female rat reproductive cyclicity and hormone secretion.

BACKGROUND: Ethanol exposure impairs mammalian reproductive function. However, the mechanisms are not fully understood. METHODS: Adult female rats were given an ethanol or a calorically matched control diet. A third group was given a liquid nonethanol diet. Half the animals were killed at 2 weeks (short chronic) and the other half at 2 months (long chronic), all on the day of proestrous on the basis of daily vaginal smears. RESULTS: The major effect of ethanol feeding was disruption in the estrous cycle. Although all of the pair-fed animals continued to cycle, 40% of the ethanol rats in the short chronic study had disruption of their cycles. In the long chronic study, 83% of the ethanol animals had abnormal cycling, in contrast to 16% of the pair-fed controls. The nature of the cycle disruption was prolongation of diestrous, with an increased time interval between proestrous surges. In four ethanol-fed rats, there was complete cessation of the estrous cycle. However, ethanol did not decrease ovarian or uterine weight. Ethanol significantly increased serum estradiol in the short chronic but not long chronic study, whereas progesterone was unchanged. Ethanol did cause a significant reduction in circulating insulin-like growth factor. CONCLUSIONS: The major effect of both short chronic and long chronic ethanol exposure was disruption of the estrous regularity, leading to a decreased number of proestrous surges. Part of the mechanism of this disruption might be a transient estrogen increase or a decrease in circulating insulin-like growth factor.

Peripubertal paternal EtOH exposure.

Fetal alcohol syndrome usually implies effects on the offspring of maternal EtOH consumption during gestation, with fewer reports addressing the impact of paternal exposure on the progeny. One previous report has dealt with the impact of EtOH exposure on peripubertal male rats as a model of teenage drinking and the deleterious effects on the offspring. We report here findings examining the effect of 2 mo of EtOH feeding on male animals as they progressed through puberty on their ability to impregnate EtOH-naive female rats and characteristics of the subsequent litters. The EtOH-imbibing fathers weighed significantly less than pairfed controls and animals ingesting a non-EtOH liquid diet ad libitum. Nevertheless, they were able to mate successfully, although fecundity was significantly reduced. The number of successful pregnancies, defined as carried to term, was diminished from 92% in controls to 75% in EtOH-fed animals (p < 0.05). There was increased paternal testicular oxidative injury demonstrated by enhanced lipid peroxidation, protein oxidation, and decreased ratio of reduced to oxidized glutathione. The litter size of the EtOH-exposed males was reduced by 46%. The average litter size was 12.4+/-1.5 pups/litter in ad libitum animals, virtually identical to the 12.5+/-0.6 pups/litter in the pair fed controls. This is in sharp contrast to the 6.7+/-0.1 pups/litter from the paternal EtOH matings (p < 0.001). There was an increase in the average individual weight of pup offspring of paternally EtOH-exposed animals (p < 0.01 vs pair-fed controls and p < 0.05 vs ad libitum). Curiously, the male-to-female pup ratio was altered with a higher preponderance of male offspring from EtOH-fed fathers. There were no gross malformations noted among the pups. Insulin-like growth factor-1 levels in the pups at 10 d of age were unaltered between the groups. However, leptin was significantly elevated in the EtOH offspring. It appears that chronic EtOH exposure in the peripubertal fathers subsequently decreases fecundity and that this may be mediated by testicular oxidative injury, perhaps leading to accelerated germ cell apoptosis.

Ethanol-induced hypogonadism is not dependent on activation of the hypothalamic-pituitary-adrenal axis.

The well-characterized suppression of the male reproductive unit after ethanol (EtOH) exposure has been speculated to be partially due to activation of the hypothalamic-pituitary-adrenal (HPA) axis. The subsequent corticosterone elevation could result in hypogonadism via suppression of hypothalamic LHRH, pituitary LH, or a direct gonadal effect. To directly examine this possibility, adult male Sprague Dawley rats were either adrenalectomized (ADX) or sham ADX. The ADX animals were given low dose corticosterone via replacement pellet, resulting in a steady level of serum corticosterone. The sham ADX (adrenal intact) animals were implanted with placebo pellets. Half of both groups were then exposed to EtOH by I.P. injection on two consecutive days to mimic an acute binge-drinking model. The other half was given saline I.P., serving as controls. In the adrenal intact animals, EtOH caused the expected rise in corticosterone, and fall in luteinizing hormone (LH) and testosterone. In the ADX animals, where constant levels of corticosterone were maintained by pellet implantation, EtOH resulted in similar LH and testosterone reduction. These results suggest that suppression of the reproductive axis is independent of the activation of the HPA unit.

Impact of acute and chronic ethanol exposure on prolactin in both male and female rats.

The deleterious effects of ethanol (EtOH) on reproduction have been well documented. This disruption is usually associated with alterations in prolactin (PRL) levels, which is relevant since this hormone is an important participant in the reproductive system. Reported EtOH-induced changes in PRL (i.e., stimulation or inhibition) have varied. These differences may have been owing to the gender or age/sexual maturity of the animal and the mode of the administration of EtOH. Therefore, to clarify the impact of EtOH on PRL, a series of experiments were conducted utilizing rats of both genders, exposed to EtOH acutely or chronically, as adults and as they progressed through puberty. In general, in younger animals of both genders, EtOH depressed serum PRL whether given acutely or chronically. In adult males, acute EtOH actually stimulated PRL levels while chronic administration had no effect. In adult females, EtOH's effect was highly dependent on the stage of the estrous cycle in which EtOH was given and during which PRL was measured. In conclusion, our studies have shown that the PRL response to EtOH is dependent on the gender and age/sexual maturity of the animals as well as on the mode of administration.

Ethanol exposure enhances apoptosis within the testes.

BACKGROUND: Chronic ethanol abuse causes testicular atrophy and male infertility in alcoholic men. It is well known that ethanol exposure disrupts the hypothalamic-pituitary-gonadal axis, adversely affects the secretory function of Sertoli cells, and produces oxidative stress within the testes. It is still not clear what cellular mechanisms are responsible for the morphologic alteration of the testes that results in a reduction of testicular mass as a consequence of ethanol exposure. The hypothesis tested was that ethanol enhances apoptosis of testicular germ cells. METHODS: In the experiments of chronic ethanol exposure, male Sprague Dawley rats (Harlan Sprague Dawley, Inc., Indianapolis, IN) were fed Liber-Decarlie liquid diet for 9 weeks. In the experiments of acute ethanol exposure, a small volume of 20% ethanol solution was administered by intratesticular injection. Both 3'-end labeling of isolated testicular deoxyribonucleic acid (DNA) and labeling of apoptotic cells in situ by the terminal deoxynucleotidyl transferase-mediated deoxyuridine 5'-triphosphate nick end-labeling method were used to determine apoptosis rates within the testes. The expression of proteins involved in apoptosis was assessed by reverse transcription-polymerase chain reaction and by Western blotting. RESULTS: The testes of rats that were fed an ethanol-containing liquid diet had more testicular DNA fragmentation than did those of animals that were fed an isocaloric control diet. Ethanol increased the number of apoptotic spermatogonia as well as spermatocytes. Direct intratesticular injections of ethanol solution enhanced testicular DNA fragmentation, suggesting an increase in apoptosis. Moreover, Fas ligand levels were increased within the testes of rats that were chronically fed ethanol. In vitro, ethanol treatment of cultured Sertoli cells enhanced the production of Fas ligand. In addition, testicular levels of p53 messenger ribonucleic acid were increased in rats that were chronically fed ethanol. CONCLUSIONS: All of these observations suggest that ethanol enhances testicular germ cell apoptosis.

Effect of intensive glycemic control on microalbuminuria in type 2 diabetes. Veterans Affairs Cooperative Study on Glycemic Control and Complications in Type 2 Diabetes Feasibility Trial Investigators.

OBJECTIVE: Microalbuminuria can reflect the progress of microvascular complications and may be predictive of macrovascular disease in type 2 diabetes. The effect of intensive glycemic control on microalbuminuria in patients in the U.S. who have had type 2 diabetes for several years has not previously been evaluated. RESEARCH DESIGN AND METHODS: We randomly assigned 153 male patients to either intensive treatment (INT) (goal HbA(1c) 7.1%) or to standard treatment (ST) (goal HbA(1c) 9.1%; P = 0.001), and data were obtained during a 2-year period. Mean duration of known diabetes was 8 years, mean age of the patients was 60 years, and patients were well matched at baseline. We obtained 3-h urine samples for each patient at baseline and annually and defined microalbuminuria as an albumin:creatinine ratio of 0.03-0.30. All patients were treated with insulin and received instructions regarding diet and exercise. Hypertension and dyslipidemia were treated with similar goals in each group. RESULTS: A total of 38% of patients had microalbuminuria at entry and were evenly assigned to both treatment groups. INT retarded the progression of microalbuminuria during the 2-year period: the changes in albumin:creatinine ratio from baseline to 2 years of INT versus ST were 0.045 vs. 0.141, respectively (P = 0.046). Retardation of progressive urinary albumin excretion was most pronounced in those patients who entered the study with microalbuminuria and were randomized to INT. Patients entering with microalbuminuria had a deterioration in creatinine clearance at 2 years regardless of the intensity of glycemic control. In the group entering without microalbuminuria, the subgroup receiving ST had a lower percentage of patients with a macrovascular event (17%) than the subgroup receiving INT (36%) (P = 0.03). Use of ACE inhibitors or calcium-channel blockers was similarly distributed among the groups. CONCLUSIONS: Intensive glycemic control retards microalbuminuria in patients who have had type 2 diabetes for several years but may not lessen the progressive deterioration of glomerular function. Increases in macrovascular event rates in the subgroup entering without albuminuria who received INT remain unexplained but could reflect early worsening, as observed with microvascular disease in the Diabetes Control and Complications Trial.

Influence of sex hormonal status on alcohol-induced oxidative injury in male and female rat liver.

BACKGROUND: Oxidative stress contributes to the development of liver injury after chronic alcohol intake. Women exhibit greater sensitivity to alcohol-induced liver disease than do men. The aim of the study was to determine the relationship between the sex hormone status of male and female rats and the degree of alcohol-induced oxidative stress in the liver. METHODS: Male and female rats were pair-fed a liquid diet that contained 36% of their total daily calories as ethanol (EtOH group) or maltose (control group). Blood and liver samples were collected at the end of 8 weeks of diet. RESULTS: Male EtOH rats experienced a reduction in plasma testosterone (T) and an increase in estradiol (E2) levels, with an increase in their calculated E2/T ratio with respect to their controls. Malonaldehyde (MDA) levels, an index of lipid peroxidation, and protein carbonyl content, an index of protein oxidation, in the liver were greater among the EtOH groups in females than in males. In males, an inverse correlation was found between hepatic MDA and circulating T levels, and a direct correlation was disclosed between MDA and estradiol levels. In addition, the hepatic histopathological score correlated inversely with the plasma T levels and directly with the calculated E2/T ratio, an index of feminization. CONCLUSIONS: Alcohol-induced oxidative injury, which contributes to hepatic injury in both male and female rats, is enhanced in females compared with males. A role for plasma T levels in protecting male rat liver from ethanol-induced oxidative injury can be hypothesized.

Two years of intensive glycemic control and left ventricular function in the Veterans Affairs Cooperative Study in Type 2 Diabetes Mellitus (VA CSDM).

OBJECTIVE: The Veterans Affairs Cooperative Study in Type 2 Diabetes Mellitus (VA CSDM) was a multicenter randomized prospective study of 153 male type 2 diabetic patients to assess the ability to sustain clinically significant glycemic separation between intensive and standard treatment arms. A trend toward an excess of combined cardiovascular events in the intensive treatment arm of this trial was reported earlier. The present analysis was done to evaluate the effect of 2 years of intensive glycemic control on the left ventricular (LV) function. RESEARCH DESIGN AND METHODS: The patients were randomized to intensive step treatment with insulin alone or with sulfonylurea (intensive treatment arm [INT], n = 75) or to standard once-daily insulin injection (standard treatment arm [STD], n = 78) treatment. A total of 136 patients (standard treatment arm [STD], n = 70; INT, n = 66) had radionuclide ventriculography at entry and at 24 months for the assessment of LV function. RESULTS: There was no difference in the mean LV ejection fraction (at entry: STD 57.1+/-9.51%; INT 58.1+/-8.7%; at 24 months: STD 57.3+/-10.8%, INT 59.5+/-10.7%), peak filling rate (at entry: STD 2.6+/-0.7 end diastolic volume per second, INT 2.4+/-0.8 end diastolic volume per second; at 24 months: STD 2.7+/-1.0 end diastolic volume per second, INT 2.5+/-0.7 end diastolic volume per second), or time to peak filling rate (at entry: STD 195.3+/-69.5 ms, INT 185.6 +/-62.4 ms; at 24 months: STD 182.6+/-64.8 ms, INT 179.2+/-61.2 ms) between the 2 treatment arms. A subgroup analysis of 104 patients (STD, n = 53; INT, n = 51) that omitted individuals with intervening cardiac events/revascularization or a change in cardioactive medications also showed no difference in the LV function at entry and at 24 months between the 2 groups. Abnormal LV ejection fraction at baseline predicted cardiac events (interval between cardiac beats [RR] = 2.5). CONCLUSIONS: Two years of intensive glycemic control does not affect the LV systolic or diastolic function in patients with type 2 diabetes.

Chronic alcohol consumption during male rat adolescence impairs skeletal development through effects on osteoblast gene expression, bone mineral density, and bone strength.

BACKGROUND: The effect of chronic alcohol ingestion on bone formation is mediated through its direct actions on osteoblasts. The affected population of mature osteoblasts declines in both number and function resulting in decreased cancellous bone volume and cortical bone strength. Although the mechanism of action on osteoblasts is unknown, alcohol alters osteoblast gene expression and matrix synthesis. METHODS: Male rats consuming alcohol (EtOH) daily for 60 days from 35 days of age until 95 days of age (unrecovered group) were compared to rats switched to a regular diet of rat chow without EtOH for an additional 90 days (recovered group). The effects of chronic dietary EtOH on skeletal development during adolescence were examined in the unrecovered and recovered rats by hormonal analysis, bone mineral density determination, bone histomorphometry, metaphyseal gene expression for osteoblast-specific proteins, and biomechanical analysis. RESULTS: The unrecovered EtOH imbibing rats weighed less than their paired isocaloric-fed and ad libitum mates. Statistically significant reductions occurred in femur lengths in the unrecovered EtOH-fed group compared to controls. Serum testosterone levels were significantly decreased by EtOH consumption but returned to higher normal levels during the recovery period. Serum insulin-like growth factor-1 (IGF-1) levels were unaffected by EtOH. Serum osteocalcin levels in the unrecovered EtOH-fed group were higher than those in the recovered group but EtOH intake did not elevate the unrecovered levels compared to isocaloric or ad libitum control rats. Quantitative computed tomography (QCT) determination of bone mineral density (BMD) revealed a statistically significant reduction only in the distal femur metaphysis in the unrecovered EtOH-fed rats. BMD increased during recovery in the distal femur metaphysis and femur mid-cortex. Image analysis of midsagittal sections of the proximal tibial metaphysis of unrecovered rats revealed reductions in cancellous area, trabecular cellularity and thickness, and increased trabecular separation. Cortical widths were significantly reduced by chronic EtOH consumption. These changes remained statistically significant at the end of the recovery period. Four-point biomechanical testing of femurs from EtOH-fed and control unrecovered groups revealed significant reductions in cortical strength, energy-to-failure, and stiffness. These cortical characteristics returned to normal values with abstinence. Tibial metaphyseal alpha-1 type I collagen and osteocalcin mRNA expression levels were significantly elevated above the paired isocaloric control levels after 60 days of EtOH consumption. Metaphyseal alkaline phosphatase mRNA levels remained unaltered by EtOH consumption in the unrecovered group. After 90 days of abstinence alpha-1 type I collagen and alkaline phosphatase gene expression levels remained significantly elevated over the isocaloric and ad libitum control levels (collagen) and the isocaloric control value (alkaline phosphatase). However, metaphyseal osteocalcin mRNA levels declined to normal levels during abstinence. CONCLUSIONS: Chronic consumption of EtOH during the peripubertal period of skeletal growth leads directly to decreased metaphyseal and cortical bone mediated through effects on osteoblasts. Removal of EtOH from the diet is accompanied by incomplete restoration of normal bone metabolism during skeletal growth.

Effects of Nitric oxide synthase blockade on the acute response of the reproductive axis to ethanol in pubertal male rats.

The effects of ethanol (EtOH) and nitric oxide (NO) are well known in the adult male rat reproductive axis. In the present study, we investigate the effects of EtOH, NO, and their interaction on key genes and reproductive hormone levels in mid- (45-day) and late pubertal (55-day) male rats. Using three different NO synthase blockers--N'omega-nitro-L-arginine methyl ester (L-NAME), N'omega-nitro-L-arginine (L-NA), and 7-nitroindazole--we show that it is possible to block, in part, some of the disruptive effects of EtOH. L-NAME totally prevented the EtOH-induced fall in serum testosterone in both 45- and 55-day-old rats (p < 0.05 and p < 0.001, respectively). On the other hand, the D-NAME, an inactive isomer of L-NAME, did not protect testosterone from suppression caused by EtOH. Similarly, L-NA and 7-nitroindazole prevented the suppression of testosterone caused by EtOH in 55-day-old animals (p < 0.001 L-NA and p < 0.05 for 7-nitroindazole), but not in the 45-day-old rats. Serum luteinizing hormone (LH) was significantly reduced by EtOH in all the studies in both age groups. L-NAME (but not D-NAME) and L-NA prevented this inhibition in 55-day-old animals (p < 0.001 for L-NAME and p < 0.01 for L-NA). However, only L-NA was able to prevent the effects of EtOH on LH in the 45-day-old rats. 7-Nitroindazole was unable to prevent the decrease in LH in either age group. Despite changes in the other reproductive hormones, there were no consistent changes in hypothalamic concentrations of either LH releasing hormone (LHRH) or its precursor, pro-LHRH. No treatment caused any change in steady-state levels of beta-LH mRNA. There were no consistent changes in pro-LHRH mRNA; but, interestingly, in 45-day-old rats, L-NA given with or without EtOH lead to a significant fall in LHRH gene expression. Our findings indicate that the acute suppressive effects of EtOH on the hypothalamic-pituitary-gonadal axis of the pubertal male rat can be at least partially prevented by NO synthase blockade.

Reversal of chronic ethanol-induced testosterone suppression in peripubertal male rats by opiate blockade.

Teenage drinking continues to be a significant problem in the U.S., as well as abroad. We have previously demonstrated that opiate blockade with naltrexone, a drug currently used in patients to diminish alcohol craving, prevented the fall in serum testosterone seen after acute ethanol (EtOH) exposure in young, peripubertal male rats. To follow-up on this reversal, a series of experiments was performed to determine if naltrexone would also prevent the testosterone suppression caused by chronic EtOH exposure. Peripubertal rats either 45 days old (mid-pubertal) or 55 days old (late pubertal) were fed an EtOH-containing liquid diet or pair-fed control diet for 14 days. Each animal was implanted with either a naltrexone containing or placebo pellet before starting the liquid diet. In each age group, EtOH alone significantly suppressed testosterone, whereas naltrexone prevented this fall, although it had no effect alone. Serum luteinizing hormone was also suppressed by EtOH; however, naltrexone did not abrogate this fall. In the 45-day-old animals, beta-luteinizing hormone mRNA levels rose significantly in the EtOH group, but not when naltrexone was coadministered with EtOH. There was no change in hypothalamic luteinizing hormone releasing hormone (LHRH) mRNA, pro-LHRH, or LHRH in any group at either age. Thus, naltrexone is able to partially prevent the EtOH-induced suppression of gonadal testosterone of young, adolescent male rats. This effect appears to be mediated directly at gonadal level, because hypothalamic and pituitary hormone changes were minor and nonsignificant.

Impact and reversibility of chronic ethanol feeding on the reproductive axis in the peripubertal male rat.

Teenage drinking continues to be a major problem in the United States as well as abroad. A significant depression in serum testosterone in adolescents who consume EtOH has been well described. In the male rodent model, a similar fall in testosterone has been reported, and prevention with the opiate blocker naltrexone has been demonstrated. To explore further the impact of chronic EtOH exposure on the reproductive axis in peripubertal rats, we designed this study specifically to define whether or not there was recovery after abstinence by examining reproductive hormones and their genes during and after EtOH exposure. Peripubertal male rats 35 d old were fed an EtOH-containing diet or a calorically matched control diet for 60 d. A third group was fed the control liquid diet ab libitum. EtOH was then withdrawn and all animals were fed standard rat chow and water ad libitum for an additional 3 mo. The EtOH-imbibing animals were found consistently to weigh less than their pair-fed mates and liquid diet ad libitum animals. Serum testosterone levels and testicular weights were significantly decreased by EtOH whereas serum estradiol levels were higher, suggesting enhanced peripheral conversion by EtOH. Spermatogenesis, assessed by histological parameters, was unaltered by EtOH. Serum luteinizing hormone levels were not different among the groups. Hypothalamic luteinizing hormone-releasing hormone mRNA levels were unaffected by EtOH. During the 3-mo recovery period, all the changes reversed, with a significant increase noted in testosterone. All other parameters remained the same among the groups. Thus, although chronic EtOH exposure in the peripubertal age period results in significant reproductive alterations, there is complete recovery on withdrawal.

The effects of intensive glycemic control on neuropathy in the VA cooperative study on type II diabetes mellitus (VA CSDM).

To determine whether a difference in HbA(1c) could be safely sustained between a standard therapy (STD) arm and an intensive therapy (INT) arm, while maintaining HbA(1c) levels in both arms within a range acceptable in community practice. The effects of intensive treatment on various parameters were studied in this feasibility trial. We report here the results of 24 months of INT on peripheral and autonomic neuropathy.A prospective trial was conducted in five medical centers in 153 men of 60 +/- 6 years of age who had a known diagnosis of diabetes for 7.8 +/- 4 years. They were randomly assigned to a standard insulin treatment group (one morning injection per day) or to an intensive therapy group designed to attain near-normal glycemia and a clinically significant separation of glycohemoglobin from the standard arm. A four-step plan was used in the intensive therapy group along with daily self-monitoring of glucose: (1) an evening insulin injection, (2) the same injection adding daytime glipizide, (3) two injections of insulin alone, and (4) multiple daily injections. Peripheral neuropathy was diagnosed clinically by a history and physical examination, and by abnormal autonomic neuropathy Valsalva ratio (VR < 1.2) and RR variation (RRV < 10). An average HbA(1c) separation of 2.07% was achieved with INT, having HbA(1c) at or below 7.3% (p = 0. 001 versus STD). Baseline prevalence of peripheral neuropathy was 53% in STD, and 48% in INT. By 24 months, the prevalence increased to 69% in STD (p = 0.005 versus baseline), and to 64% in INT (p = 0. 008 versus baseline, but no different than STD). Though INT did not reverse all elements of peripheral neuropathy, there was a decreased prevalence of cranial neuropathy (p = 0.053 versus STD) and more frequent preservation of touch sensation in the upper extremities (p = 0.03 versus STD) in INT. At baseline, an abnormal Valsalva ratio and/or RR variation was seen in 38% of STD and 31% of INT. By 24 months in STD, the prevalence rose to 55% (p = 0.0067 versus baseline), and in INT, to 48% (p = 0.012 versus baseline and no different from STD). The prevalence of erectile dysfunction increased from 53% at baseline to 73% at 2 years, p = 0.002 in STD, and from 51% to 73% at 2 years (p = 0.003 versus baseline) and no different from STD. There was no change in the frequency of abnormal gastrointestinal or sweating symptoms. Our conclusion was that 2 years of meticulous glycemic control did not decrease overall prevalence of peripheral or autonomic neuropathy. In fact, the prevalence rose equivalently and significantly in both treatment arms. There was some benefit, however, in decreased frequency of cranial neuropathy and better preservation of touch sensation in INT.

Effect of nitric oxide synthase inhibitors on preventing ethanol-induced suppression of the hypothalamic-pituitary-gonadal axis in the male rat.

Ethanol (EtOH) suppression of the hypothalamic-pituitary-gonadal (HPG) axis results in broad reproductive malfunction. In the HPG axis, the suppressive effects of EtOH are manifested by decreased serum testosterone, reduced testicular luteinizing hormone (LH) receptor numbers, lowered serum LH and pituitary beta-LH mRNA levels (in castrated animals), and impaired luteinizing hormone releasing hormone (LHRH) release from the hypothalamus. Increasing evidence has suggested that nitric oxide (NO) plays a role in regulation of the HPG axis. NO was shown to stimulate LHRH secretion from the hypothalamus and to have variable effects on LH release from the pituitary. At the gonadal level, NO is inhibitory to testosterone production. NO may directly inhibit some testicular steroidogenic enzymes. To investigate the effect of EtOH, NO, and their interaction on the male HPG axis, three NO synthase (NOS) inhibitors, N(G)-nitro-L-arginine methyl ester, N(G)-nitro-L-arginine, and 7-nitro indazole were used to study overall HPG function in the presence and absence of EtOH. Animals were given intraperitoneal injections of saline, EtOH, various NOS inhibitors, or EtOH, along with NOS inhibitors 2 hr before sacrifice. Serum testosterone and LH concentrations, pituitary beta-LH mRNA levels, hypothalamic LHRH mRNA levels, and LHRH content were determined. It was found that blocking NOS by these NOS inhibitors prevented EtOH-induced suppression of testosterone and, in some cases, serum LH. However, this was not accompanied by concurrent changes with NOS blockade on LHRH mRNA, hypothalamic pro-LHRH or LHRH content or pituitary LH beta mRNA levels. It appears that the protective effect of NOS blockade was largely, although not completely, due to a direct effect at the gonadal level.

Interaction of ethanol and nitric oxide in the hypothalamic-pituitary-gonadal axis in the male rat.

Ethanol (EtOH) exerts deleterious actions on reproductive function at all three levels: the hypothalamus, pituitary, and gonad (HPG). Nitric oxide (NO), a newly identified messenger molecular in a variety of biological systems, has been suggested as playing a role in HPG hormone regulation. NO stimulates luteinizing hormone releasing hormone secretion from the hypothalamus and has variable effects on luteinizing hormone release from the pituitary. NO is inhibitory to testosterone production, and it may also directly inhibit some steroidogenic enzymes. Related studies in the accompanying paper have demonstrated that inhibiting NO synthase (NOS) using various NOS inhibitors can prevent the EtOH-induced suppression of testosterone on the male HPG axis, and this action is mainly, although not entirely, due to a direct gonadal effect. To further investigate the role of NO in the HPG axis, we assessed the HPG NO-NOS system by determining NOS mRNA levels, protein levels, and enzyme activity in the presence and absence of EtOH. At the testicular level, EtOH's action did not appear to be mediated by increasing NO content. However, EtOH was able to potentiate NO's suppressive effect on the testicular synthesis system. One locus where EtOH and NO interacted was at the steroidogenic enzyme level. N(G)-nitro-L-arginine methyl ester, a NOS inhibitor, was found to antagonize the EtOH-induced fall on P-450 17alpha-hydroxylase/C17-20 lyase mRNA levels when administered along with EtOH. EtOH had no apparent effect on the pituitary NO-NOS system and its effects on the hypothalamic NO-NOS system do not explain its ability to reduce luteinizing hormone releasing hormone secretion.

Reversal of ethanol-induced testosterone suppression in peripubertal male rats by opiate blockade.

Teenage drinking is a major problem in the United States, as well as abroad. Besides psychosocial implications, ethanol (EtOH) has detrimental effects on the reproductive system. Clinical problems associated with reduced reproductive hormones include osteoporosis, decreased muscle function, anemia, altered immune function, prostate involution, and decreased reproductive abilities. Education coupled with strategies aimed at preventing these deleterious consequences even in the face of continued EtOH intake is extremely important. We have tested the possibility that naltrexone, a drug currently used in patients to decrease alcohol craving, might also prevent the fall in the male hormone, testosterone, caused by EtOH exposure. Rats aged 35 days old (prepubertal), 45 days old (midpubertal), and 55 days old (late pubertal) were injected (intraperitoneally) with either saline, EtOH, naltrexone, or EtOH plus naltrexone. In the two older age groups, EtOH significantly suppressed testosterone, which was prevented by administration of naltrexone. In the youngest animals, there was no treatment effect presumably due to low basal levels of testosterone. EtOH similarly reduced luteinizing hormone (LH), but this suppression was not prevented by naltrexone. There was no consistent effect of any treatment on hypothalamic concentration of pro-LH releasing hormone (RH) (LHRH), LHRH, or on steady-state levels of LHRH mRNA. We conclude that, as animals progress through puberty, EtOH suppresses LH and testosterone. The testosterone decline can be prevented by opiate blockade with naltrexone, an effect primarily seen at gonadal level. Thus, naltrexone, a drug already used clinically to reduce EtOH intake, also has protective physiological effects on the endocrine system.

The role of gonadectomy and testosterone replacement on thymic luteinizing hormone-releasing hormone production.

We and others have identified luteinizing hormone-releasing hormone (LHRH) in cells of the immune system in both animals and humans. LHRH is an immunostimulant, and testosterone is an immunosuppressant. Because testosterone is known to modulate the concentrations of hypothalamic LHRH, we wondered whether testosterone might also alter the concentrations of rat thymic LHRH. Two weeks after castration or sham castration, adult male rats were implanted with either vehicle or testosterone capsules. All animals were killed 4 days after capsule implantation. Thymic LHRH concentration increased significantly in castrated animals. Testosterone replacement prevented this increase. The concentration of the LHRH precursor, proLHRH, decreased significantly, but testosterone replacement prevented this decrease. Steady-state concentrations of LHRH mRNA were not changed by castration or by hormonal replacement. In contrast to the post-castration increase in thymic LHRH, LHRH content of the hypothalamus decreased significantly. Whereas concentrations of LHRH were lower in the thymus than in the hypothalamus, proLHRH concentrations were much greater in the thymus. These data suggest that gonadal manipulation modulates LHRH molecular processing and its tissue concentration in the thymus in addition to those in the hypothalamus, and that the regulation of LHRH molecular processing by testosterone in the hypothalamus is different from that in the thymus.