Factors influencing required dietary nutrient concentrations.

A review of the genetic and environmental factors that must be considered when forming nutritionally adequate diets for specific laboratory animal colonies.

Nutrition of rodents.

Access to a diet that provides adequate nutrition is one of the most important environmental factors influencing the well-being of rodent colonies. The dietary ingredient and nutrient composition, as well as the potential biological and chemical contaminant concentrations, are factors for consideration in selecting diets for a specific rodent colony. Estimated nutrient requirements have been published for the rodent species that are commonly used in biomedical research. The nutrient concentrations in adequate diets for other captive rodent species that are not used in biomedical research are more difficult to obtain. However, reasonable estimates of their nutrient requirements can be obtained by extrapolation of data from rodent species of a similar metabolic weight and size or from nutrient concentrations of diets that have a history of acceptable performance in the species of interest. Captive rodent colonies should be provided with nutritionally balanced diets with only limited amounts of succulent foodstuffs. The practice of feeding rodent colonies specific cereal grains is discouraged, since no single grain provides a balanced rodent diet.

Sugar-induced blood pressure elevations over the lifespan of three substrains of Wistar rats.

OBJECTIVE: Since the majority of studies concerned with sugar-induced blood pressure elevation have principally been short-term, the present investigation followed the effects of heavy sucrose ingestion on systolic blood pressure (SBP) and related parameters over the lifespan of three substrains of Wistar rats. METHODS: Two hundred twenty-five rats (75 spontaneously hypertensive rats (SHR), 75 Wistar Kyoto rats (WKY), 75 Munich Wistar rats (WAM) were given one of five diets. The baseline diet in terms of calories derived 32% from sucrose, 33% from protein, and 35% from fat. The remaining four diets derived their calories as follows: a high sugar-low protein diet--52% of calories from sucrose, 15% from protein, and 33% from fat; a high sugar-low fat diet--53% of calories from sucrose, 37% from protein, and 10% from fat; a low sugar-high protein diet--11% calories from sucrose, 56% from protein, and 33% from fat, and a low sugar-high fat--13% of calories from sucrose, 32% from protein, and 55% from fat. RESULTS: All substrains showed the highest systolic blood pressure when ingesting the two diets highest in sucrose. The highest sugar-induced SBP elevation, which remained over the lifespan of all substrains, was found in SHR. WKY had an intermediate elevation. WAM showed the lowest responses, although the average elevation of 6-8 mm Hg was statistically significant. The following parameters could not be correlated with long-term elevation of SBP; body weight, catecholamine excretion, renal function, and plasma renin activity. Only insulin concentrations correlated: insulin concentrations were consistently higher in the two groups of WKY and WAM consuming the high sucrose diets. CONCLUSIONS: High dietary sucrose can chronically increase SBP in three substrains of Wistar rats. Increased concentrations of circulating insulin were found in WKY and WAM suggesting that the glucose/insulin system was involved, at least in these two substrains, in the maintenance of high SBP levels during chronic, heavy sugar ingestion.

Dietary restriction slows age pigment accumulation in the retinal pigment epithelium.

PURPOSE: The accumulation of age pigment, or lipofuscin, in postmitotic cells appears to be a universal feature of the aging process in animals. In mammals, the lipofuscin content of the retinal pigment epithelium (RPE) increases progressively during senescence. Dietary restriction has been shown to slow the rate at which many biologic parameters change during aging. Experiments were conducted to determine if dietary restriction alters the rate of age pigment accumulation in the RPE. METHODS: Male Wistar rats were placed on one of three dietary regimens starting at weaning. One group was fed a nutritionally complete diet ad libitum. Another group was fed the same diet but was only allowed to consume 60% as much food daily as the ad libitum group ate. The final group was fed ad libitum a nutritionally complete diet that had a lower caloric density per gram than the diets fed to the other animals primarily because of the replacement of carbohydrate with oat fiber. Ultrastructural morphometric analysis was used to determine the RPE age pigment content in the first group at 6 months of age, and in all of the groups at 18 months of age. RESULTS: Dietary restriction, achieved either by reducing total food intake or by reducing the caloric content of the diet, resulted in significant decreases in RPE lipofuscin accumulation. CONCLUSIONS: Dietary restriction provides a relatively simple means by which RPE age pigment content can be modulated. This should prove useful in assessing the role of RPE lipofuscin accumulation in age-related retinal disorders. That the oat fiber diet fed ad libitum was almost as effective as restriction of total food intake in slowing RPE age pigment accumulation indicates that the effect of restricted caloric intake is not mediated by almost constant hunger.

Dietary restriction in nonhuman primates: progress report on the NIA study.

Rhesus and squirrel monkeys have been fed a semisynthetic diet at approximately ad libitum or 30% reduced levels for 3.5 (rhesus group 2) to 4.5 (rhesus group 1 and squirrel) years. Animals have maintained excellent health status as determined by physical examinations, hematology, and blood chemistry. While relative rates of body weight gain in restricted group 1 rhesus and squirrel monkeys have been markedly reduced, DR effects on crown-rump length (body height) have been variable. In addition, numerous physiological and biochemical parameters have been measured, and several exhibit significant cross-sectional age effects. Interestingly, several of these also exhibit possible species and genotype (group 1 and 2 rhesus) differences. A number of physiological parameters are emerging that might be altered by DR; however, further explanation of these effects awaits more extensive and detailed analyses.

High sucrose diets increase blood pressure of both salt-sensitive and salt-resistant rats.

We examined the effects of a diet relatively high in sugar and low in protein content on systolic blood pressure (SBP) in rats with known pressure responses to salt (NaCl) in order to compare "sugar/protein sensitivity" to "salt sensitivity." Dahl salt-sensitive (DSS) and salt-resistant (DSR) rats were fed one of two low salt diets containing either high sugar (sucrose 51.5% w/w)/low protein (14.6% w/w) or low sugar (sucrose 12.5% w/w)/high protein (52.2% w/w) content. After 3 weeks, the DSS ingesting the high sugar diet/low protein diet developed significantly elevated SBP relative to DSR eating the same high sugar/low protein diet and the DSS and DSR consuming the low sugar/high protein diet. After 2 to 3 months, the SBP of DSR eating the high sugar diet began to rise markedly and eventually both DSS and DSR ingesting the high sugar/low protein diet maintained similarly elevated SBP, significantly higher than DSS and DSR ingesting the low sugar/high protein diet. When Fischer 344 rats, a normotensive, salt-resistant rat strain, were fed the high sucrose/low protein diet, SBP also rose significantly into hypertensive ranges over 2 to 3 months. Since the SBP of DSR and Fischer 344 rats are not influenced to any great extent by high salt intake, even after prolonged exposure, the SBP rise associated with the high sugar/low protein diet may be via a mechanism different from salt-induced hypertension. However, it is also possible that the high sugar/low protein diet creates in DSS and DSR the situation responsible for salt induction in DSS.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood pressure responses to sucrose ingestion in four rat strains.

Among four strains examined, spontaneously hypertensive rats (SHR) show a marked (20 mm Hg, P less than .01) systolic blood pressure elevation (SBP), Sprague-Dawley (SD) and Wistar-Kyoto (WKY) rats developed a moderate elevation (8 mm Hg, P less than .01), and a normotensive Wistar rat (WAM) had a lesser SBP elevation (6 mm Hg, P = NS) after excess sucrose ingestion. The SBP elevations found in SHR were noted at 2 and 4 weeks after starting the dietary treatments. Corresponding with SBP changes, plasma renin activity (PRA), aldosterone, and neuropeptide Y (NPY) concentrations all decreased with the high sucrose-low protein diet compared to the low sucrose-high protein diet, while circulating insulin levels were unchanged. Although norepinephrine (NE) and epinephrine (E) excretion tended to be higher in the rats eating the high sucrose-low protein food, the differences were not statistically significant. The differences in these parameters could influence the SBP in SHR, SD, and WKY, but virtually similar qualitative and quantitative blood and urinary findings were found in WAM, a strain of rat that showed no significantly increased SBP. Removing one kidney increases the CHO-induced SBP response of WKY to levels comparable to those seen in SHR, converting a moderate responder to a highly sensitive one. We conclude that under well-controlled conditions there are obvious differences in the SBP response to the macronutrients in the diets of various rat strains and that SHR possess some intrinsic mechanism(s), most likely associated with renal metabolism, which make this strain more sensitive to refine CHO-induced SBP elevations.

High-moisture diet for laboratory rats: nutrient analysis, growth, and organ weights.

A diet (KSC-25) to be sterilized by irradiation was formulated to contain 66% moisture and to provide the required nutrients for growing rats. Analyses of the irradiated dry diet provided data to evaluate its nutrient content. The diet was evaluated for its ability to supply all nutrients, including water, required by immature rats. Sixteen Sprague-Dawley rats were fed the high-moisture diet with or without access to a water bottle. Rats (n = 16) fed an irradiated purified diet in a meal form with access to a water bottle were the control animals. Feed efficiency, food and water consumption, and growth rate data were collected during the 28-day study. Organ weights were collected on day 28. The test diet met or exceeded the National Research Council (NRC) estimated nutritional requirements for immature laboratory rats. The 66% moisture KSC-25 diet provided all nutrients, including water, required by weanling male Sprague-Dawley rats for growth equivalent to the established purified diet.

High-moisture diet for laboratory rats: complete blood counts, serum biochemical values, and intestinal enzyme activity.

Rats were fed an irradiated high-moisture diet (KSC-25) with or without access to a water bottle. Physiologic values were compared between these two groups and a group of rats fed a purified diet. Hematologic and serum biochemical values, urine specific gravity, and intestinal enzyme activities were determined from samples collected from the three groups of rats. Sprague Dawley rats (n = 32) fed the irradiated high-moisture diet with or without a water bottle were the test animals. Rats (n = 16) fed an irradiated purified diet and water provided via a water bottle were the control group. The purified diet formulation modified AIN-76A, is a commonly used purified diet for laboratory rodents. All rats remained alert and healthy throughout the study. A comparison of the physiologic values of rats in this study with reported normal values indicated that all of the rats in the study were in good health. Significant differences (P less than 0.05) of the physiologic values from each rat group are reported.

Development of sugar-induced blood pressure elevation after uninephrectomy in a resistant rat strain.

Certain rat strains acutely increase blood pressure (BP) when given diets high in NaCl. Prior results showed that "salt-sensitive" rat strains, at least the ones studied, also increase BP in response to sugar loading. To examine this relationship further and learn more about the pathogenesis of sucrose-induced BP elevations, we examined the effects of unilateral nephrectomy (uninephrectomy) on sucrose-induced BP changes. The rationale is based upon the findings that renal mass removal sensitizes BP response to salt loading. Over 15 weeks, augmented sugar (sucrose) consumption by Long-Evans (LE) rats did not increase BP markedly compared to rats consuming a diet relatively low in sugar unless uninephrectomy was performed. The differences in BP caused by the high sugar diet in a uninephrectomized rat could not be explained adequately by alterations in catecholamine excretion, plasma renin activity, excesses in blood volume, or the other parameters examined. However, salt-induced hypertension has been attributed to the presence of circulating substances affecting ion transport. Among the dietary groups, there was a significant correlation between the ability of plasma to depress PAH and TEA renal slice uptake and the difference in BP. This is consistent with the presence of a circulating factor affecting cell transport that has its greatest activity in the high sugar-uninephrectomy group of LE rats. We conclude that reducing renal mass potentiates sugar-induced BP elevation similar to salt-induced BP elevation in a normally resistant rat strain, and the rise of BP may be caused by a circulating factor.

Hemorrhagic cardiomyopathy and hemothorax in vitamin K deficient mice.

The cause of a fatal condition characterized by hemorrhagic cardiomyopathy, hemothorax, and coagulation defects in hysterectomy-derived male mice was investigated. Microscopic heart alterations included multifocal hemorrhage and necrosis with variable degrees of acute inflammation and fibroplasia that were most severe in the region of the atrioventricular junction. A spontaneous outbreak was arrested by increasing menadione Na-bisulfite (vitamin K) in the feed to 20 ppm. The complete syndrome including hemorrhagic cardiomyopathy was readily reproduced in germ-free male mice given a vitamin K-free diet, and in conventional male and female mice given Warfarin in the diet. We concluded that the cause of this condition was vitamin K deficiency.

Prevention of cataracts in pink-eyed RCS rats by dark rearing.

Royal College of Surgeons rats have hereditary retinal degeneration and associated posterior subcapsular opacities (PSO) of the lens, detectable by slitlamp at 7-8 postnatal weeks in both pink- and black-eyed rats. The retinal degeneration is intensified by light, especially in pink-eyed rats. A fourth of pink-eyed rats developed mature cataracts by 9-12 months of age, but black-eyed rats whose retinas are protected from light by pigmented irises and pigment epithelium rarely have mature cataracts (3% or less), indicating light may be a factor in cataractogenesis. Prior work had shown that dark rearing reduced the rate of retinal degeneration in pink- but not black-eyed rats, but cataracts were not studied. In the present work, pregnant pink-eyed females were placed in a darkroom 1 week before parturition. Pups were removed over intervals at 20-85 postnatal days for: (a) microscopic study of fresh lenses and of fixed, stained retina and lens, and (b) counts of cells mm-2 of the web-like vitreous cortex after it had been dissected free. The macrophage-like cells are a quantitative index of immune reaction to retinal damage. At 50-53 postnatal days, in pink-eyed cyclic light reared RCS, the mean number of macrophages was 4.6-fold that in congenic controls, but in those that were dark reared it was only 1.4-fold. This was less than the increase in cyclic light reared black-eyed RCS (2.3-fold that in congenic black-eyed controls). Total absence of light reduced retinal degeneration and the number of macrophages, and prevented PSO detectable microscopically.(ABSTRACT TRUNCATED AT 250 WORDS)

Dietary restriction and aging: the initiation of a primate study.

Juvenile (1 yr) and adult (3-5 yr) male rhesus monkeys (Macaca mulatta) and juvenile (1-4 yr) and adult (5-10 yr) male squirrel monkeys (Saimiri sciureus) were fed a diet at or near ad libitum levels based on recommended caloric intake for age and body weight or fed 30% less of the same diet with this restriction gradually introduced over a 3-mo period. Analysis of body weights among these respective control and experimental groups from the first year of the study indicated that the monkeys undergoing dietary restriction were gaining weight at a markedly slower rate compared to control values. Actual food intake among diet-restricted groups had been reduced 22-24% below control levels. Periodic analysis of hematology and blood chemistry measurements over the first year of the study detected few significant differences between control and experimental groups to indicate that diet restriction was not detrimental to general health. When values obtained from hematology and blood chemistry measurements of juvenile and adult groups (control and experimental groups combined) were compared to ad libitum fed old monkeys from each species (greater than 18 yr for rhesus; greater than 10 yr for squirrel monkeys), many significant age differences were noted. Among the largest and most consistent findings in both species were age-related decreases in concentrations of lymphocytes, serum glutamic oxalacetic transaminase, serum glutamic pyruvic transaminase, alkaline phosphatase, and phosphates as well as the albumin/globulin ratio and the blood urea nitrogen/creatinine ratio. Age-related increases in serum globulin and creatinine concentrations were also found. These parameters as well as many others being implemented in the study will be monitored further to determine if diet restriction affects the rate of development as well as aging as observed in numerous rodent studies applying such nutritional manipulations.

Excess sucrose and glucose ingestion acutely elevate blood pressure in spontaneously hypertensive rats.

Spontaneously hypertensive rats (SHR) were fed 6 different diets. The baseline diet (I) derived equal calories from sucrose, proteins, and fats. Three other diets (II, III, VI) derived the majority of calories from refined CHO, sucrose or glucose, with decreases in calories from proteins or fats. The last two diets (IV, V) were relatively low in sucrose with a higher percentage of the total calories from proteins and fats, respectively. From 3 to 15 weeks on the diets, the highest average BP was in rats consuming high concentrations of sucrose or glucose (II, III, VI). Urinary excretory rates of norepinephrine (NE) at 5, 10 and 15 weeks and epinephrine at 5 and 10 weeks were significantly elevated in rats ingesting diets high in refined CHO, and NE positively correlated with blood pressure (BP) at 5 and 10 weeks of the study. At the end of the study, serum insulin levels were not different, but plasma renin and serum glucagon levels were lower in SHR consuming the diets with high CHO concentrations. We conclude that equally elevated BP are seen with relatively high intakes of either sucrose or glucose, whether the balance of calories is derived from lessening fat or protein. This is secondary, at least in part, to alterations in NE metabolism.

Dietary trans fatty acids modulate erythrocyte membrane fatty acyl composition and insulin binding in monkeys.

The substitution of trans- for half of the cis-monounsaturated fatty acids in the diet of Macaca fasicularis monkeys resulted in alterations in erythrocyte fatty acid composition and insulin receptor properties but not in membrane fluidity. Both cis and trans diets contained 10% fat and similar fatty acid compositions, except that approximately 50% of the cis-octadecenoate (c-18:1) in the cis diet was replaced with trans-octadecenoate isomers (t-18:1) in the trans diet. Compared with the cis diet, the trans diet resulted in the incorporation of approximately 11% t-18:1, an approximately 50% decrease in c-18:1, an approximately 16% decrease in total saturated fatty acids, and an approximately 20% increase in 18:2(n-6) in erythrocyte membrane lipids. The increase in 18:2(n-6) may reflect on homeostatic mechanisms designed to maintain overall membrane fluidity, as no diet-related changes in fluidity were observed with diphenylhexatriene steady state fluorescence polarization. Values observed for insulin binding and insulin receptor number were higher and binding affinity was lower in monkeys fed the cis diet. In the absence of an effect on overall membrane fluidity, altered receptor activity suggests that insulin receptor activity is dynamic, requiring specific fluid membrane subdomains or highly specific fatty acid-protein interactions.

Effect of sodium and potassium ingestion on renal growth in rats.

To determine whether differences in dietary Na and K intake influence renal growth and compensatory renal growth following unilateral nephrectomy (uni), rats were given 2 diets for 2 weeks which differed only in Na-K concentrations. Diet 1 (High Na, Low K) contained 1.0% Na, 0.36% K and diet 2 (Low Na, High K) contained 0.05% Na, 2.0% K. Half of the rats consuming diets 1 and 2 for 1 week received uni and the other half of sham operation (sham). The rats were followed for another week while consuming the experimental diets. Uni and sham rats on diet 1 showed significant increase in blood pressure (BP) (136 +/- 4 v 126 +/- 3 mm Hg, P less than .05). Uni itself did not alter blood pressure. Body weight and heart rate were unchanged by diet or operation. Kidney weight, renal RNA, protein, RNA/DNA, and protein/DNA increased significantly after uni but the differences were not affected by diet. Serum blood urea nitrogen (BUN) and creatinine were not markedly different among any group of rats suggesting no major renal damage. We conclude that dietary changes of Na-K which cause no obvious renal damage even though BP is elevated moderately do not influence renal growth or compensatory renal growth. Based on RNA, DNA and protein metabolism, the form of growth (hypertrophy or hyperplasia) is also not influenced by renal electrolyte handling.

Influence of oat bran on sucrose-induced blood pressure elevations in SHR.

To determine whether oat fiber influences BP, we gave spontaneously hypertensive rats (SHR) a diet high in sucrose and low in protein (calories: sucrose 52%, protein 15%, fat 33%) or a diet low in sucrose and high in protein (calories: sucrose 13%, protein 52%, fat 35%). The amount of fat in these particular diets has not been shown to influence BP, so we modified the 2 diets by replacing fat with oat bran (10% w/w). Accordingly, we examined 4 groups of 5 rats consuming different diets: high sucrose, high sucrose + oat bran, low sucrose, and low sucrose + oat bran. Not unexpectedly, SHR consuming the diet high in sucrose had a significantly higher BP after 2 weeks than those consuming the diet low in sucrose. The significant difference in BP continued over the next 3 weeks. At the end of 6 week duration of study, we found the following BP: SHR ingesting the high sucrose diet, 217 mm Hg +/- 5 (SEM) vs SHR consuming the low sucrose diet, 187 mm Hg +/- 4 (SEM) p less than .0001]. SHR eating the low sucrose diet and consuming supplemental bran showed no significant change in BP after 6 weeks compared to SHR eating the basic diet alone, 188 mm Hg +/- 6 (SEM); however, 5 SHR consuming the high sucrose diet with added oat bran showed a significantly lower BP 200 mm Hg +/- 2 (SEM) than SHR ingesting the basic high sucrose diet devoid of oat bran [p less than .01]. We conclude that addition of oat bran to the diet can ameliorate sucrose-induced BP elevations in SHR.

Maintenance of cotton-top tamarins fed an experimental pelleted diet versus a highly diverse sweetened diet.

The future study of colon disease in captive callitrichid colonies may require manipulation of diets. The limited knowledge of the nutritional requirements for these species and the varied diets and supplementations fed to these animals in various colonies suggest the importance of testing the palatability and acceptability of diets for these primates. Individually housed cotton-top tamarins (Saguinus oedipus) were given either the regular Oak Ridge Associated Universities (ORAU) diet (monkey chow slurry, canned diet and supplements), a similar slurry using an experimental natural ingredient diet plus supplements, or the experimental diet without supplements. Neither dry food consumption, body weight, fecal output, nor the histological evaluation of the colons were affected by these diets. Daily intake of protein and calories were higher than previously reported estimates for the species. These results demonstrate that a natural ingredient non-sweetened pelleted diet is palatable for cotton-top tamarins for a period of 3.5 months, however, further testing over longer time periods is necessary. The nonnutritional (e.g. psychological) advantages of providing a highly diverse diet to primates housed in a relatively monotonous environment should be considered before adopting such a diet for an entire colony.

The apparent reversal of a wasting syndrome by nutritional intervention in Saguinus mystax.

One hundred eighty sexually mature Saguinus mystax were imported from Peru in six lots over a period of 1 year. Within 1 year after arrival, the mortality was 60% and the majority of the tamarins showed signs similar to "wasting marmoset syndrome" (WMS). In an effort to improve the survival rate, an open formula diet replaced the commercial closed formula diet that had been fed since arrival of the tamarins. The open formula diet contained 26.2% crude protein, 12.3% ether extract, 43.3% nitrogen free extract and 5.9% crude fiber on a dry matter basis. The diet was evaluated on the basis of palatability, weight gain, mortality, digestibility, nitrogen balance, serum biochemical parameters and blood counts. The mean daily consumption on an as-is basis was 44.8g or 335 Kcal gross energy/Kg of body wt./day. During the 3 month open formula diet evaluation period average weight increased by 56g (p less than .05), mortality decreased demonstratively, and alopecia and chronic diarrhea were nearly eliminated. Mean daily gross energy intake for S. mystax (335 Kcal/Kg of body wt/day) was substantially greater than previously reported values for callitrichids. WMS signs observed in the S. mystax colony were controlled by providing what appears to be an adequate diet.

Contaminant and nutrient concentrations of natural ingredient rat and mouse diet used in chemical toxicology studies.

The NIH-07 open formula natural ingredient rat and mouse ration is the standard diet for chemical toxicity and carcinogenicity studies conducted for the National Toxicology Program (NTP). Contaminant and nutrient concentrations were determined in 2 to 94 lots of this diet used in the NTP toxicology studies. All nutrient concentrations were equivalent to or greater than the requirements for rats and mice as set forth by the National Research Council. Aflatoxins, Hg, chlorinated hydrocarbons except methoxychlor, organophosphates except malathion, estrogenic activity, and Salmonella sp. were not present at the detectable levels. Fluorine, As, Cd, Pb, Se, N-nitrosodimethylamine, N-nitrosopyrrolidine, N-nitrosomorpholine, nitrate, nitrite, butylated hydroxyanisole, butylated hydroxytoluene, ethylene dibromide, methoxychlor, malathion, and trypsin inhibitor activity were present at or above the detectable levels. Five lots of diet had nitrosamine content of 100 to 273 ppb and 7 lots had 2.08 to 3.37 ppm of Pb. All other lots of NIH-07 diet used for NTP toxicology studies contained low levels of the contaminants. After determination of the contaminant concentrations in the 94 lots of diet and the contaminant concentrations in natural ingredients used in formulating NIH-07 diet, maximum allowable levels of contaminants were established and a flexible scoring system for acceptability of each lot of diet for chemical toxicology studies was developed. By prescreening ingredients such as fish meal for heavy metals and nitrosamines, and applying the flexible scoring system proposed, more than 95% of the lots of NIH-07 diet produced during the last 3 years had scores of greater than or equal to 95 out of 100 points and were considered acceptable for toxicology studies.