Effect of Isoflurane Anesthesia on Circadian Metabolism and Physiology in Rats.

Isoflurane anesthesia alters the blood levels of several neuroendocrine hormones associated with normal metabolism and physiology and increases stress, but the effect of brief CO2 anesthesia on these parameters is unknown. In this study, we examined the effects of isoflurane (4%) compared with brief CO2 (70% CO2, 30% air) anesthesia on circadian rhythms of plasma measures of physiology and metabolism. Adult male Sprague-Dawley rats (Crl:SD; n = 6 per group) were maintained on a 12:12-h light:dark (300 lx; lights on, 0600) photoperiod. After 1 wk of acclimation, a series of 6 low-volume blood draws were collected by cardiocentesis under anesthesia using isoflurane (10 min or less) compared with CO2 (1 min or less) at a single circadian time point every 4 d (0400, 0800, 1200, 1600, 2000, or 2400) over 3 wk to assess arterial blood glucose, lactic acid, and potassium and plasma melatonin, leptin, insulin, total fatty acids, and corticosterone concentrations. Results revealed that plasma levels (mean ± SEM) of melatonin were low (11 ± 1 pg/mL) during the light phase in both groups but were significantly lower during the dark phase in the isoflurane group (48 ± 6 pg/mL) compared with the CO2 group (162 ± 18 pg/mL). In addition, prominent circadian rhythms of arterial plasma levels of corticosterone, glucose, total fatty acids, lactic acid, and potassium were altered in the isoflurane group compared with the CO2 group. These findings demonstrate that the normal circadian rhythms of endocrine physiology and metabolism observed during brief CO2 anesthesia in rats are markedly disrupted by isoflurane anesthesia.

Effects of Colored Enrichment Devices on Circadian Metabolism and Physiology in Male Sprague-Dawley Rats.

Environmental enrichment (EE) gives laboratory animals opportunities to engage in species-specific behaviors. However, the effects of EE devices on normal physiology and scientific outcomes must be evaluated. We hypothesized that the spectral transmittance (color) of light to which rats are exposed when inside colored enrichment devices (CED) affects the circadian rhythms of various plasma markers. Pair-housed male Crl:SD rats were maintained in ventilated racks under a 12:12-h light:dark environment (265.0 lx; lights on, 0600); room lighting intensity and schedule remained constant throughout the study. Treatment groups of 6 subjects were exposed for 25 d to a colored enrichment tunnel: amber, red, clear, or opaque. We measured the proportion of time rats spent inside their CED. Blood was collected at 0400, 0800, 1200, 1600, 2000, and 2400 and analyzed for plasma melatonin, total fatty acids, and corticosterone. Rats spent more time in amber, red, and opaque CED than in clear tunnels. All tubes were used significantly less after blood draws had started, except for the clear tunnel, which showed no change in use from before blood sampling began. Normal peak nighttime melatonin concentrations showed significant disruption in the opaque CED group. Food and water intakes and body weight change in rats with red-tinted CED and total fatty acid concentrations in the opaque CED group differed from those in other groups. These results demonstrate that the color of CED altered normal circadian rhythms of plasma measures of metabolism and physiology in rats and therefore might influence the outcomes of scientific investigations.

Effect of different spectral transmittances through tinted animal cages on circadian metabolism and physiology in Sprague-Dawley rats.

The suprachiasmatic nucleus is synchronized by the light:dark cycle and is the master biologic clock that serves as a pacemaker to regulate circadian rhythms. We explored the hypothesis that spectral transmittance (tint) of light through caging alters circadian rhythms of endocrine and metabolic plasma constituents in nonpigmented Sprague-Dawley rats. Rats (Crl:SD; n = 12 per group) were housed in a 12:12-h light:dark environment (300 lx; 123.0 µ W/cm(2); lights on, 0600) in either clear-, amber-, blue-, or red-tinted rodent cages. Blood was collected at 0400, 0800, 1200, 1600, 2000, and 2400 and measured for melatonin, total fatty acids, pH, glucose, lactic acid, corticosterone, insulin, and leptin. As expected, plasma melatonin levels were low during the light phase but higher during the dark phase in all groups; however, when compared with the clear-cage group, rats in amber-, blue-, and red-tinted cages had 29%, 74%, and 48%, respectively, greater total daily melatonin levels due to an increased duration and, in some cases, amplitude of the nocturnal melatonin signal. No differences were found in dietary and water intake, body growth rates, total fatty acids, pH, or glucose among groups. Disruptions in circadian rhythms, manifesting as alterations in phase timing, amplitude, or duration, occurred in the melatonin, lactic acid, corticosterone, insulin, and leptin levels of rats in tinted compared with clear cages. Therefore, the use of variously tinted animal cages significantly alters circadian rhythms in plasma measures of metabolism and physiology in laboratory rats, thus potentially altering the outcomes of scientific investigations.

Effect of spectral transmittance through red-tinted rodent cages on circadian metabolism and physiology in nude rats.

Light entrains normal circadian rhythms of physiology and metabolism in all mammals. Previous studies from our laboratory demonstrated that spectral transmittance (color) of light passing through cages affects these responses in rats. Here, we addressed the hypothesis that red tint alters the circadian nocturnal melatonin signal and circadian oscillation of other metabolic and physiologic functions. Female nude rats (Hsd:RH-Foxn1(rnu); n = 12 per group) were maintained on a 12:12-h light (300 lx; 123.0 µW/cm(2); lights on 0600):dark regimen in standard polycarbonate translucent clear or red-tinted cages. After 1 wk, rats underwent 6 low-volume blood draws via cardiocentesis over a 4-wk period. Plasma melatonin levels were low during the light phase (1.0 ± 0.2 pg/mL) in rats in both types of cages but were significantly lower in red-tinted (105.0 ± 2.4 pg/mL) compared with clear (154.8 ± 3.8 pg/mL) cages during the dark. Normal circadian rhythm of plasma total fatty acid was identical between groups. Although phase relationships of circadian rhythms in glucose, lactic acid, pO2, and pCO2 were identical between groups, the levels of these analytes were lower in rats in red-tinted compared with clear cages. Circadian rhythms of plasma corticosterone, insulin, and leptin were altered in terms of phasing, amplitude, and duration in rats in red-tinted compared with clear cages. These findings indicate that spectral transmittance through red-colored cages significantly affects circadian regulation of neuroendocrine, metabolic, and physiologic parameters, potentially influencing both laboratory animal health and wellbeing and scientific outcomes.

The surface glycoprotein of feline leukemia virus isolate FeLV-945 is a determinant of altered pathogenesis in the presence or absence of the unique viral long terminal repeat.

Feline leukemia virus (FeLV) is a naturally transmitted gammaretrovirus that infects domestic cats. FeLV-945, the predominant isolate associated with non-T-cell disease in a natural cohort, is a member of FeLV subgroup A but differs in sequence from the FeLV-A prototype, FeLV-A/61E, in the surface glycoprotein (SU) and long terminal repeat (LTR). Substitution of the FeLV-945 LTR into FeLV-A/61E resulted in pathogenesis indistinguishable from that of FeLV-A/61E, namely, thymic lymphoma of T-cell origin. In contrast, substitution of both FeLV-945 LTR and SU into FeLV-A/61E resulted in multicentric lymphoma of non-T-cell origin. These results implicated the FeLV-945 SU as a determinant of pathogenic spectrum. The present study was undertaken to test the hypothesis that FeLV-945 SU can act in the absence of other unique sequence elements of FeLV-945 to determine the disease spectrum. Substitution of FeLV-A/61E SU with that of FeLV-945 altered the clinical presentation and resulted in tumors that demonstrated expression of CD45R in the presence or absence of CD3. Despite the evident expression of CD45R, a typical B-cell marker, T-cell receptor beta (TCRß) gene rearrangement indicated a T-cell origin. Tumor cells were detectable in bone marrow and blood at earlier times during the disease process, and the predominant SU genes from proviruses integrated in tumor DNA carried markers of genetic recombination. The findings demonstrate that FeLV-945 SU alters pathogenesis, although incompletely, in the absence of FeLV-945 LTR. Evidence demonstrates that FeLV-945 SU and LTR are required together to fully recapitulate the distinctive non-T-cell disease outcome seen in the natural cohort.

Effects of spectral transmittance through standard laboratory cages on circadian metabolism and physiology in nude rats.

Light is potent in circadian, neuroendocrine, and neurobehavioral regulation, thereby having profound influence on the health and wellbeing of all mammals, including laboratory animals. We hypothesized that the spectral quality of light transmitted through colored compared with clear standard rodent cages alters circadian production of melatonin and temporal coordination of normal metabolic and physiologic activities. Female nude rats (Hsd:RH-Foxn1(rnu); n = 6 per group) were maintained on a 12:12-h light:dark regimen (300 lx; lights on, 0600) in standard translucent clear, amber, or blue rodent cages; intensity and duration of lighting were identical for all groups. Rats were assessed for arterial blood levels of pO(2) and pCO(2), melatonin, total fatty acid, glucose, lactic acid, insulin, leptin, and corticosterone concentrations at 6 circadian time points. Normal circadian rhythms of arterial blood pO(2) and pCO(2) were different in rats housed in cages that were blue compared with amber or clear. Plasma melatonin levels (mean ± 1 SD) were low (1.0 ± 0.2 pg/mL) during the light phase in all groups but higher at nighttime in rats in blue cages (928.2 ± 39.5 pg/mL) compared with amber (256.8 ± 6.6 pg/mL) and clear (154.8 ± 9.3 pg/mL) cages. Plasma daily rhythms of total fatty acid, glucose, lactic acid, leptin, insulin, and corticosterone were disrupted in rats housed in blue or amber compared with clear cages. Temporal coordination of circadian rhythms of physiology and metabolism can be altered markedly by changes in the spectral quality of light transmitted through colored standard rodent cages.

Eliminating animal facility light-at-night contamination and its effect on circadian regulation of rodent physiology, tumor growth, and metabolism: a challenge in the relocation of a cancer research laboratory.

Appropriate laboratory animal facility lighting and lighting protocols are essential for maintaining the health and wellbeing of laboratory animals and ensuring the credible outcome of scientific investigations. Our recent experience in relocating to a new laboratory facility illustrates the importance of these considerations. Previous studies in our laboratory demonstrated that animal room contamination with light-at-night (LAN) of as little as 0.2 lx at rodent eye level during an otherwise normal dark-phase disrupted host circadian rhythms and stimulated the metabolism and proliferation of human cancer xenografts in rats. Here we examined how simple improvements in facility design at our new location completely eliminated dark-phase LAN contamination and restored normal circadian rhythms in nontumor-bearing rats and normal tumor metabolism and growth in host rats bearing tissue-isolated MCF7(SR(-)) human breast tumor xenografts or 7288CTC rodent hepatomas. Reducing LAN contamination in the animal quarters from 24.5 ± 2.5 lx to nondetectable levels (complete darkness) restored normal circadian regulation of rodent arterial blood melatonin, glucose, total fatty and linoleic acid concentrations, tumor uptake of O(2), glucose, total fatty acid and CO(2) production and tumor levels of cAMP, triglycerides, free fatty acids, phospholipids, and cholesterol esters, as well as extracellular-signal-regulated kinase, mitogen-activated protein kinase, serine-threonine protein kinase, glycogen synthase kinase 3ß, γ-histone 2AX, and proliferating cell nuclear antigen.

Concentration and emission of airborne contaminants in a laboratory animal facility housing rabbits.

Characterization of animal housing conditions can determine the frequency of bedding and cage changes, which are not standardized from facility to facility. Rabbits produce noticeable odors, and their excreta can scald and stain cages. Our facility wanted to document measurable airborne contaminants in a laboratory rabbit room in which excreta pans were changed weekly and cages changed biweekly. Contaminants included particulate, endotoxin, ammonia, carbon dioxide, and a rabbit salivary protein as a marker for rabbit allergen. Concentrations were measured daily over a 2-wk period in a laboratory animal facility to determine whether they increased over time and on days considered to be the dirtiest. Except for ammonia, concentrations of all airborne contaminants did not differ between clean and dirty days. Concentrations were lower than occupational health exposure guidelines for all contaminants studied, including ammonia. After measurement of concentration, a model was applied to calculate mean emission factors in this rabbit room. Examples of emission factor utilization to determine airborne contaminant concentration in rabbit rooms under various environmental conditions and housing densities are provided.

Complications of gastric catheters implanted in rhesus macaques (Macaca mulatta).

As part of a study addressing chronic alcohol consumption and simian immunodeficiency virus, 31 rhesus macaques (Macaca mulatta) were implanted with gastric catheters used to deliver alcohol or isocaloric sucrose (control). Once implanted, the animals wore jackets and were housed in specialized cages modified with swivels and tethers. During the course of the study, 3 animals developed clinical signs indicating possible instability of the implanted gastric catheter. All 3 animals were found to have a string foreign body wrapped around the distal end of the catheter, with 2 of the catheters perforating the intestinal wall. Gastroscopy was used to screen remaining animals to determine catheter position and the presence of a foreign body attached to the end of the catheter. Results of the screening revealed that of the 28 remaining animals, 9 had malpositioned catheters; string foreign bodies were associated with 3 of the 9 malpositioned catheters. We initially hypothesized that the peristaltic motion of the stomach, combined with the attachment of string, which was probably ingested by subjects after manipulating their jackets, led to eventual catheter displacement. We later concluded that the string may have played a secondary role but was not the primary cause of catheter instability, because several malpositioned catheters had no string attached at the time of diagnosis. Subsequent modifications were instituted, including modifying the surgical technique, altering the type of gastric catheter used, and increasing environmental enrichment for animals with known tendency to manipulate their jackets.

Clinical and hematological effects of serial phlebotomy performed on laboratory beagles.

We conducted a study designed to mimic a typical pharmacokinetic study to gain a better understanding of a dog's response to multiple, frequent blood sampling at 15% total blood volume. Ten dogs were randomly assigned to either a control group having sham venipuncture performed or to a blood collection group having 1.5% of their body weight (approximately 15% total blood volume) removed weekly for 4 weeks. Both groups were monitored during a 2-week recovery period immediately after the 4-week collection period. Parameters evaluated were clinical signs, body weight, and hematological and serum biochemical analytes. There were minimal differences in red blood cell morphology between the two groups. Statistically significant differences in hematocrit between the two groups occurred on several days, and this finding was attributed to blood withdrawal in the blood collection group; however, this statistical difference was not deemed to be clinically significant. There were no statistically significant differences in body weight, total protein, reticulocyte count, mean corpuscular volume, mean corpuscular hemoglobin, or red cell distribution width. We conclude that removing 15% blood volume in laboratory beagles is compatible with maintaining the health and well-being of the dog and can be acceptable in laboratory situations when it is scientifically justified.