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Behavioral, physical, and demographic changes in Drosophila populations through dietary restriction.

Dietary restriction (DR) is a valuable experimental tool for studying the aging process. Primary advancement of research in this area has relied on rodent models, but attention has recently turned toward Drosophila melanogaster. However, little is known about the baseline effects of DR on wild-type Drosophila and continued experimentation requires such information. The findings described here survey the effects of DR on inbred, wild-type populations of Canton-S fruit flies and demonstrate a robust effect of diet on longevity. Over a circumscribed range of dietary conditions, healthy lifespan varies by as much as 121% for wild-type Drosophila females. Significant differences are also observed for male flies, but the magnitude of the DR effect is less robust. Mortality analyses of the survivorship data reveal that this variation in lifespan can be attributed to a modulation of the rate parameter for the mortality function - a change in the demographic rate of aging. Since the feeding of fruit flies is less easily controlled than that of rodents, this research also addresses the validity of applying a DR model to Drosophila populations. Feeding and body weight data for flies given the various dietary conditions surveyed indicate that Drosophila on higher-calorie diets consume a similar volume of food to those on a low-calorie diet, resulting in different levels of calorie intake. Fertility and activity levels demonstrate that the diets surveyed are comparable, and that increasing the calorie content of laboratory food up to twice the normal concentration is not pathologic for experimental fly populations.

Aging↗

Plasma leptin is partly cleared by the kidney and is elevated in hemodialysis patients.

Leptin, the gene product of the ob gene, is important in the control of appetite in rodents and may have an important role in humans. The clearance of leptin from the circulation is unknown. As the leptin receptor is present in the kidney, we evaluated the role of the kidney in removing circulating leptin in humans. We measured leptin in aortic and renal vein plasma in 8 patients with intact renal function and 6 patients with impaired renal function who were undergoing elective cardiac catheterization. Renal blood flow was measured in all patients to calculate net mass balance across the kidney. In patients with intact renal function there is net renal uptake of 12% of circulating leptin, whereas in patients with renal insufficiency there is no renal uptake of leptin. In a separate cohort of 36 patients with end-stage renal failure on hemodialysis, peripheral leptin levels factored for body mass index was increased by > fourfold as compared to a group of healthy controls (N = 338). In addition, plasma leptin is not cleared by hemodialysis with a modified cellulose membrane. Additional studies are required to evaluate the role of leptin in mediating the anorexia of uremia.

Aged↗

Effects of three intravenous lipid emulsions on the survival and mononuclear phagocyte function of septic rats.

The immunosuppressive effects of intravenous lipid emulsions are a matter of great concern and debate. In a rat model of gram-negative bacteremia, we assessed whether the use of three intravenous lipid emulsions with different triacylglycerol compositions could influence mortality, bacterial clearance, and prostaglandin E(2) (PGE(2)) levels and compared these groups with groups of orally fed rats and rats that received a small amount of calories in form of glucose without enteral feeding (starvation). RATS WERE ASSIGNED TO ONE OF FIVE GROUPS: group 1 (control, n = 15) received rodent chow ad libitum and saline infusion; group 2 (starvation group, n = 12) had no access to chow and received an infusion of 5% glucose; group 3 (n = 17) received total parenteral nutrition (TPN) with long-chain triacylglycerols; group 4 (n = 12) received TPN with medium- and long-chain triacylglycerols; and group 5 (n = 15) received TPN with its emulsion based on olive oil. Animals received isonitrogenous and isocaloric TPN. After 2 d of TPN, a dose of 10(8) colony-forming units of Escherichia coli was introduced via the venous catheter; 2 d later the animals were killed. Blood, spleen, liver, and lungs were cultured. Circulating levels of PGE(2) were measured. Bacterial growth in the liver and lungs were significantly higher in groups 3 and 4 than in group 1, with no differences among the other groups. Rates of bacteremia were significantly higher in groups 3 and 4 than in group 1, with no differences among the other groups. Plasma levels of PGE(2) did not differ, and mortality was unaffected. Bacterial clearance clearly was preserved in orally fed, control rats when compared with rats on TPN with long-chain triacylglycerols or medium- plus long-chain triacylglycerols. However, the use of a lipid emulsion enriched intravenously with oleic acid was a valid way of reducing this disturbance, although plasma levels of PGE(2) and survival were not modified.

Administration, Oral↗

Dichlorvos toxicity in the white-footed mouse (Peromyscus leucopus).

The organophosphate pesticide, dichlorvos (DDVP), is used commonly to control ectoparasites in laboratory rodents colonies. This compound is relatively nontoxic to Mus musculus at dosages several times the therapeutic level. However, usage of a similar therapeutic level in the white-footed mouse (Peromyscus leucopus) resulted in substantial mortality. To determine whether P. leucopus is more susceptible than M. musculus to the toxic effects of DDVP, both species were exposed to 0, 3 and 6 g of pelleted DDVP per cage. In a subsequent experiment, P. leucopus were exposed to 0 and 1 g of DDVP per cage. Mortality was not observed in M. musculus at any dosage level. P. leucopus exposed to 1, 3 and 6 g of DDVP exhibited mortalities of 3%, 20% and 53%, respectively. Mean serum cholinesterase in P. leucopus exposed to 3 and 6 g of DDVP was 0.35 and 0.21 U/ml as compared to 3.13 U/ml in unexposed mice. The analogous values for M. musculus were 1.60 and 0.79 U/ml while the level in unexposed mice was 6.79 U/ml. In the second experiment, mean serum cholinesterase in P. leucopus exposed to 1 g of DDVP was 0.32 U/ml as compared to 2.33 U/ml in unexposed mice. Histopathology revealed no lesions in the brain, liver or kidneys. The increased susceptibility of P. leucopus to the toxic effects of DDVP was related to the lowered serum cholinesterase. This indicates that DDVP should not be used for control of ectoparasites in P. leucopus.

Animals↗

Estradiol enhances prostaglandin E2 receptor gene expression in luteinizing hormone-releasing hormone (LHRH) neurons and facilitates the LHRH response to PGE2 by activating a glia-to-neuron signaling pathway.

Prostaglandin E2 (PGE2) mediates the stimulatory effect of norepinephrine (NE) on the secretion of luteinizing hormone-releasing hormone (LHRH), the neuropeptide controlling reproductive function. In rodents, this facilitatory effect requires previous exposure to estradiol, suggesting that the steroid affects downstream components in the cascade that leads to PGE2-induced LHRH release. Because astroglia are the predominant cell type contacting LHRH-secreting nerve terminals, we investigated the involvement of hypothalamic astrocytes in the estradiol facilitation of PGE2-induced LHRH release. A subpopulation of LHRH neurons was found to express the mRNA encoding the PGE2 receptor subtype EP1-R, which is coupled to calcium mobilization. The LHRH-producing cell line GT1-1 also contains EP1-R mRNA and, to a lesser extent, the three alternatively spliced forms of EP3-R mRNA (alpha, beta, and gamma) that encode receptors linked to inhibition and stimulation of cAMP formation. Hypothalamic astrocytes treated with estradiol produced a conditioned medium that when applied to GT1-1 cells resulted in a selective upregulation of EP1-R and EP3gamma-R mRNAs. The conditioned medium also enhanced the LHRH response to EP1-R and EP3-R agonists and the cAMP response to EP3-R activation. Thus, one mechanism by which estradiol facilitates the effect of neurotransmitters acting via PGE2 to stimulate LHRH release is by enhancing the glial production of substances that upregulate PGE2 receptors on LHRH neurons. The existence of such a mechanism underscores the emerging importance of glial-neuronal communication in the control of brain neurosecretory activity.

Animals↗

The role of small rodents and patterns of living in the epidemiology of nephropathia epidemica.

The epidemiological role of small rodents and certain human patterns of living for contraction of nephropathia epidemica (NE) was studied by interviewing 46 NE patients and 52 control subjects. The occurrence of small rodents in dwellings and cupboards was significantly commoner among NE patients than among control subjects. The results suggest a connection between NE and small rodents.

Adolescent↗

NADPH-oxidase activity and lipid peroxidation in neutrophils from rats fed fat-rich diets.

In order to investigate the effect of fat-rich diets on neutrophil functions, 21 day-aged rats were fed for 6 weeks with a control diet consisting of a regular laboratory rodent chow (4 per cent final fat content), a control diet supplied with soybean oil (15 per cent final fat content), or a control diet supplied with coconut oil (15 per cent final fat content). Glycogen-elicited peritoneal neutrophils from rats fed soybean and coconut oil-enriched diets presented a reduction in spontaneous and PMA-stimulated H2O2 generation relative to neutrophils from rats fed the control diet. The activity of superoxide dismutase, glutathione peroxidase and catalase did not change in animals fed fat-rich diets. In addition, the capacity to generate O2-, spontaneously or in response to PMA, did not change in neutrophils from animals fed fat-rich diets. Values attained matched those observed in animals fed the control diet, regardless of the method used to measure O2-, the superoxide dismutase-inhibitable reduction of cytochrome c or the lucigenin-dependent chemiluminescence. However, the initial rate of O2- generation both in resting neutrophils and in PMA-stimulated cells was significantly reduced when animals were fed with coconut or soybean oil-enriched diets due, at least in part, to a reduction in the activity of glucose-6-phosphate dehydrogenase. The concentration of thiobarbituric acid reactive substances, an index of lipid peroxidation, was increased in animals fed both fat-rich diets. This was accompanied by an increase in arachidonic acid content in these cells. Results presented suggest that lipid peroxidation in neutrophils from animals fed fat-rich diets may be associated with a consumption of H2O2 yielding more reactive oxygen-derived species such as the hydroxyl radical.

Animals↗

Do audible and ultrasonic sounds of intensities common in animal facilities affect the autonomic nervous system of rodents?

In animal facilities, noises, often poorly controlled, occur over a wide range of frequencies and intensities. Evidence demonstrates that audible noise and ultrasound have deleterious effects on rodent physiology, but it is not known how they affect the autonomic nervous system (ANS). This study exposed 3 unrestrained, male, Sprague-Dawley rats daily to a 15-min white noise regime (90 dB), a quiet regime, or a 15-min ultrasound regime (90 dB at 4 frequencies in the range 20 to 40 kHz)--each for several weeks--and used radiotelemetry to monitor their cardiovascular responses. Exposure to audible noise increased heart rate and mean arterial pressure. Spectral analysis of HR variability showed diminished stimulation of the parasympathetic nervous system, shifting the sympathovagal balance. However, ultrasound, at the frequencies used, did not reproducibly affect cardiovascular parameters. The preliminary data obtained from this study indicate that audible noise, but not ultrasound (delivered using the same protocol), affects the ANS. Because the cardiovascular, respiratory, renal, and gastrointestinal systems are under autonomic control, such noise could have wide-ranging effects on animal physiology.

Animals↗

Effects of feed restriction on fertility in female rats.

BACKGROUND: Feed restriction with its resultant body weight loss impacts the rodent estrous cycle; however, the manifestation of these changes in a regulatory study design has not been documented. This study reports the effects of feed restriction in the context of an FDA regulatory submission. METHODS: Adult female rats (n = 20/group; weighing approximately 200 g each) were provided rodent chow ad lib (control) or at 20, 15, 10, or 7.5 g/rat/day (g/day) during a 2-week pre-mating phase, throughout the mating phase, and up to gestation day (GD) 7. On GD 8, all animals were provided ad lib feed until necropsy on GD 14. Estrous cyclicity, mating, and fertility parameters were evaluated. RESULTS: Ad lib rats consumed approximately 20 and 28 g/day during the pre-mating and gestation phases, respectively. All measured fertility parameters in the 20 g/day group were similar to control values. In the 15 g/day group, body weight was reduced by 16% at 2 weeks, prolonged diestrus occurred, and fertility was compromised due to reductions in corpora lutea. Within 2 weeks, mean body weight in groups receiving < or = 10 g/day was reduced by > or = 29% compared to ad lib values, and overt changes in estrous cyclicity, mating, and fertility occurred. The 7.5 g/day group was not sustainable beyond the pre-mating phase. CONCLUSIONS: For this study type, feed intake at < or = 50% ad lib values (< or = 10 g/day) was inadequate due to the magnitude and rapidity of body weight effects. Estrous parameters appeared slightly more sensitive than functional measures, as body weight changes of approximately 16% appeared near the threshold of changing routinely calculated estrous cycle parameters and were later associated with reduced fertility. In general, body weight differences of 10-15% by themselves were not adverse to normal reproduction (20 g/day).

Animals↗

Effect of dietary treatment with n-propyl gallate or vitamin E on the survival of mice exposed to phosgene.

Phosgene, widely used in industrial processes, can cause life-threatening pulmonary edema and acute lung injury. One mechanism of protection against phosgene-induced lung injury may involve the use of antioxidants. The present study focused on dietary supplementation in mice using n-propyl gallate (nPG)--a gallate acid ester compound used in food preservation--and vitamin E. Five groups of male mice were studied: group 1, control-fed with Purina rodent chow 5002; group 2, fed 0.75% nPG (w/w) in 5002; group 3, fed 1.5% nPG (w/w) in 5002; group 4 fed 1% (w/w) vitamin E in 5002; and group 5, fed 2% (w/w) vitamin E also in 5002. Mice were fed for 23 days. On day 23 mice were exposed to 32 mg m-3 (8 ppm) phosgene for 20 min (640 mg. min m-3) in a whole-body exposure chamber. Survival rates were determined at 12 and 24 h. In mice that died within 12 h, the lungs were removed and lung wet weights, dry weights, wet/dry weight ratios, lipid peroxidation (thiobarbituric acid reactive substances, TBARS) and glutathione (GSH) were assessed. Vitamin E had no positive effect on any outcome measured. There was no significant difference between 1.5% nPG and any parameter measured or survival rate compared with 5002 + phosgene. However, dietary treatment with 0.75% nPG significantly increased survival rate (P </= 0.002) and lowered TBARS (P </= 0.05) compared with 5002 + phosgene at 12 h after exposure. Mice fed 0.75% nPG had a lower wet/dry wt ratio compared with those fed 1.5% nPG and a significantly increased lung tissue GSH 36%, compared with the 5002 + phosgene group. In conclusion, dietary treatment with a low level of the antioxidant nPG protected mice by decreasing lipid peroxidation and increasing lung tissue GSH. The higher level of nPG and both levels of vitamin E diets were ineffective, suggesting that a ceiling threshold level of antioxidants in lung tissue is required for survival against phosgene-induced lung injury. Published in 2001 by John Wiley & Sons, Ltd.

Administration, Inhalation↗

Follistatin overexpression in rodent liver tumors: a possible mechanism to overcome activin growth control.

The activin-follistatin system is a potent growth regulatory system of liver tissue homeostasis. Activin A inhibits hepatocellular DNA synthesis and induces cell death. Follistatin binds activin and sequesters it from the signaling pathway. Consistently, follistatin has been reported to act as an inducer of DNA synthesis in the liver. Using RNase protection analysis, we studied the expression of follistatin in rat and mouse liver tumors as a possible mechanism to overcome activin growth control. Approximately 40% of the tumors (nine of 24 each), most of them hepatocellular carcinomas, displayed increased levels of follistatin mRNA when compared to tumor-surrounding liver tissue. The degree of overexpression was highly variable but independent of the carcinogen treatment that animals had received. It was also independent from the histological stage of malignancy and further found in rat liver adenomas. Follistatin expression was also observed in cell lines derived from human hepatocellular carcinomas. Overexpression of follistatin may represent a unique strategy of hepatic tumors to overcome the inhibitory action of a growth factor, activin, by decreasing its local bioavailability.

Activins↗

Estradiol initially enhances but subsequently suppresses (via adrenal steroids) granule cell proliferation in the dentate gyrus of adult female rats.

In the dentate gyrus of adult female meadow voles, a high dose of estradiol benzoate (EB) increases (within 4 h) then decreases (within 48) the number of dividing progenitor cells (Ormerod BK, Galea LAM. 2001. Reproductive status regulates cell proliferation within the dentate gyrus of the adult female meadow vole: A possible regulatory role for estradiol. Neurosci 2:169-179). We investigated whether time-dependent EB exposure differentially influences the number of new granule cells produced in the adult female rat dentate gyrus and whether EB-stimulated adrenal activity mediates the decrease in cell proliferation. Ovariectomized rats received either an EB (10 microg in 0.1 mL) or vehicle (0.1 mL) injection either 4 or 48 h (Experiment 1) before a BrdU injection (200 mg/kg) and were perfused 24 h later to assess the number of new cells. Relative to vehicle, the number of new cells increased following a 4 h exposure (p < or = 0.04) but decreased following a 48 h exposure (p < or = 0.006) to EB. In Experiment 2, the number of new cells within the dentate gyrus of ovariectomized and adrenalectomized females did not significantly differ between groups exposed to EB versus vehicle for 48 h prior to BrdU administration, suggesting the decreased number of new cells observed within the dentate gyrus of adrenal-intact adult female rats is mediated by EB-stimulated adrenal activity. We conclude that estradiol dynamically regulates cell proliferation within the dentate gyrus of adult female rats in the time-dependent manner observed previously in voles and suppresses cell proliferation by influencing adrenal steroids. Investigating how estradiol dynamically regulates neurogenesis could provide insight into the mechanisms by which the proliferation of progenitor cells is controlled within the adult rodent hippocampus.

Adrenal Cortex Hormones↗

Mutation of p53 is not a prerequisite for immortalization of human fibroblasts by SV40 T antigen.

The in vitro life span of human cells is under genetic control and limited. Immortalized cells, however, can be obtained at a low frequency following expression of the SV40 T antigen gene though the steps that lead to immortality are not well understood. p53 has been implicated in cell cycle regulation and evidence suggests it may have a role in controlling life span in rodent and human cells. In this study, we investigated whether allelic loss or mutation of p53 was an essential step during SV40 immortalization leading to the appearance of immortal cell lines. The gross structure of the p53 gene was examined in a primary fibroblast cell strain (1BR.3) and two SV40-immortalized derivatives, 1BRMT1 and 1BRgn2. There was no evidence for allelic loss of the p53 gene during immortalization. The primary cells and the immortal derivatives all expressed authentic p53 mRNAs, though the immortal cell lines had higher levels of expression. Sequence analysis of exons 5-8 did not detect mutations associated with the immortal phenotype. These data are consistent with SV40 immortalization being independent of genetic changes in p53.

Antigens, Polyomavirus Transforming↗

Modulation of baseline behavior in rats by putative serotonergic agents in three ethoexperimental paradigms.

The present study adopts an ethoexperimental approach to examine the deportment subsequent to alteration in serotonin (5-HT) neurotransmission following treatment with site-specific neuropharmacological probes. The impact of perturbation in (5-HT) neurotransmission on baseline behavior was analyzed employing three animal models of anxiety, i.e., hole-board, elevated plus maze, and bright/dark arena. Inbred male rats (Wistar strain, weighing between 150 and 200 g) were used in this study. The vivarium and the behavioral laboratory were specially designed to permit operation of reversed light-dark cycle and all experiments were performed during the dark period. Pharmacological tools selected to influence 5-HT levels include (1) a combination of tranylcypromine and tryptophan (TCP + TRYPT) (0.75 mg/kg + 40 mg/kg) which augments 5-HT biosynthesis; (2) p-chlorophenylalanine (PCPA: 200 mg/kg), an inhibitor of 5-HT biosynthesis; and (3) 5-HT reuptake blockers, namely zimelidine (ZIM) (40 mg/kg) and fluoxetine (FLU) (10 mg/kg). Rats under the influence of PCPA exhibited anxiolytic response, whereas those under treatments to raise 5-HT levels, viz., TCP + TRYPT, ZIM and FLU, displayed anxiogenic-like reactions. Several other agents known to specifically interact with 5-HT receptor subtypes were also tested. 5-HT2 receptor stimulants, such as quipazine (5 mg/kg) and MK 212 (0.5 mg/kg), were found to be anxiogenic. Buspirone (2 mg/kg), a 5-HT1 agonist, surmounted normal behavioral inhibition. However, another 5-HT1 stimulant, 8-OH-DPAT (0.025 mg/kg), had anxiogenic action. Pretreatment with 5-HT3 antagonists [zacopride (2 mg/kg) and GR 38032F (0.1 mg/kg)] and putative 5-HT1 antagonist [propranolol (10 mg/kg)] resulted in borderline disinhibition of normal behavioral inhibition to novel environments. In contrast, cyproheptadine (0.5 mg/kg), a 5-HT2 antagonist, provoked anxiogenic-like behavior. Altogether, uniform results were obtained for each probe in all the three models, suggesting that the battery of anxiety tests chosen in this study is reliable and sensitive to detect unknown pharmacological responses. The results support the hypothesis that stimulation of serotonergic neurotransmission heightens normal anxiety, whereas its blockade releases normal behavioral inhibition. Furthermore, this work establishes the validity of using the three paradigms in evaluating the involvement of multiple neurotransmitter receptors in the control of behavior of rodents under natural circumstances and also detects any aberration following exposure to novelty and stress.

Animals↗

Receptor autoradiography of mu and delta opioid peptide receptors in spontaneously hypertensive rats.

The receptor autoradiographic distribution of opioid peptide receptors in spontaneously hypertensive rats (SHR) was compared to that of Sprague-Dawley (SD) rats, using the highly selective mu and delta opioid receptor ligands, [3H]DAGO (Tyr-D-Ala-Gly-NMe-Phe-Gly-ol) and [3H]DPDPE ([D-Pen2,D-Pen5]enkephalin), respectively. Although the distribution of these binding sites was similar in both strains, SHR showed significantly higher binding densities of mu receptors in 16 of 27 areas examined. These included the patch and matrix components of the caudate-putamen (CPu), olfactory tubercle, endopiriform nucleus, anterior cingulate cortex, ventral tegmental area lateroposteral thalamic nucleus and the ventral part of the dentate gyrus. In contrast, SHR had lower [3H]DAGO binding sites in the CA1 of the hippocampus. Conversely, SHR showed higher binding densities of delta receptors in 7 of 20 areas examined, including the CPu, CA2 and CA3 areas of the hippocampus and the central grey. High-to-low lateromedial gradients of striatal delta receptors were observed in both strains. Because opioid peptides are known to participate in locomotive behavior in rodents and in the control of blood pressure, the present results support a role of opioid peptidergic systems in the manifestation of hyperactivity and hypertension observed in SHR.

Animals↗

A 28-day feeding study with methyl isoeugenol in rats.

Methyl isoeugenol was administered in rodent diet for a minimum of 28 consecutive days to groups of 16 male and 16 female rats (Sprague-Dawley strain) at levels of approximately 30, 100 and 300 mg/kg body weight/day. A further group of 16 male and 16 female rats was given the rodent diet as a control. The administration of methyl isoeugenol in the diet did not adversely affect the growth or general health of the animals or their food intakes. Although high dose animals of both sexes had increased lymphocyte and total white blood cell counts, these are not considered, in isolation, to be an adverse effect of treatment. None of the minor variations observed in the serum chemical analyses or urine analyses is considered to be indicative of a treatment-related toxic effect. An increase in liver weight, adjusted for body weight, was seen in male and female rats receiving 300 mg methyl isoeugenol/kg body weight. Few histopathological abnormalities were observed. Although the incidence of kidney and Harderian gland lesions was higher for high dose animals compared with the controls, the lesions are of a type that occurs spontaneously and are thus not considered to be attributable to treatment with methyl isoeugenol. While the increased liver weight and white blood cell counts of rats given 300 mg methyl isoeugenol/kg body weight may represent effects of treatment, it is not considered that there is any reason to regard these as adverse effects.

Alanine Transaminase↗

Genetic differences in the elevated plus-maze persist after first exposure of inbred rats to the test apparatus.

The elevated plus-maze (EPM) is an anxiety model thought to assess different types of emotional states depending on whether or not the animals have been previously exposed to the test apparatus. Accordingly, benzodiazepine-treated rodents generally differ from controls in the first but not in the second EPM trial. Inbred Lewis and SHR rats of both sexes (N=10) were submitted twice (test and retest) to the EPM with a 24 h interval between trials. Overall strain differences (Lewis<SHR) were observed in both males and females concerning anxiety-related measures (time spent and percent of entries in the open arms) regardless of previous maze experience. Moreover, prior exposure to the test apparatus produced an overall decrease in the approach towards the open arms in both strains and sexes. The fact that genetic differences did not diminish or disappear in the second trial, suggests that test and retest in the EPM are likely to share some common emotional components and that differences between naïve LEW and SHR rats are not similar to those observed between control and benzodiazepine-treated animals.

Animals↗