Effect of genetic strain and gender on age-related changes in body composition of the laboratory rat.

Body fat serves as a storage compartment for lipophilic pollutants and affects the pharmacokinetics of many toxic chemicals. Understanding how body fat varies with gender, strain, and age may be essential for development of experimental models to study mechanisms of toxicity. Nuclear magnetic resonance (NMR)-based analysis serves as a noninvasive means of assessing proportions of fat, lean, and fluid in rodents over their lifetime. The aim of this study was to track changes in body composition of male and female Long-Evans (LE), Sprague-Dawley (SD), Fischer (F334), and Brown Norway (BN) rats from postweaning over a >2-yr period. Percent fat of preweaned LE and SD rats was markedly higher compared to the other strains. LE and SD strains displayed marked increases in body fat from weaning to 8 mo of age. Postweaned F344 male and females showed relatively low levels of percent fat; however, at 2 yr of age percent fat of females was equal to that of SD and LE in females. BN rats showed the highest levels of lean tissue and lowest levels of fat. Percent fat of the BN strain rose at the slowest rate as they aged. Percent fluid was consistently higher in males for all strains. Females tended to have higher percent fat than males in LE, SD, and F344 strains. Assessing changes in body fat as well as lean and fluid of various strains of male and female rats over their lifetime may prove useful in many research endeavors, including pharmacokinetics of lipophilic toxicants, mechanisms underlying obesity, and metabolic disorders.

Isolation of a Genomic Region Affecting Most Components of Metabolic Syndrome in a Chromosome-16 Congenic Rat Model.

Metabolic syndrome is a highly prevalent human disease with substantial genomic and environmental components. Previous studies indicate the presence of significant genetic determinants of several features of metabolic syndrome on rat chromosome 16 (RNO16) and the syntenic regions of human genome. We derived the SHR.BN16 congenic strain by introgression of a limited RNO16 region from the Brown Norway congenic strain (BN-Lx) into the genomic background of the spontaneously hypertensive rat (SHR) strain. We compared the morphometric, metabolic, and hemodynamic profiles of adult male SHR and SHR.BN16 rats. We also compared in silico the DNA sequences for the differential segment in the BN-Lx and SHR parental strains. SHR.BN16 congenic rats had significantly lower weight, decreased concentrations of total triglycerides and cholesterol, and improved glucose tolerance compared with SHR rats. The concentrations of insulin, free fatty acids, and adiponectin were comparable between the two strains. SHR.BN16 rats had significantly lower systolic (18-28 mmHg difference) and diastolic (10-15 mmHg difference) blood pressure throughout the experiment (repeated-measures ANOVA, P < 0.001). The differential segment spans approximately 22 Mb of the telomeric part of the short arm of RNO16. The in silico analyses revealed over 1200 DNA variants between the BN-Lx and SHR genomes in the SHR.BN16 differential segment, 44 of which lead to missense mutations, and only eight of which (in Asb14, Il17rd, Itih1, Syt15, Ercc6, RGD1564958, Tmem161a, and Gatad2a genes) are predicted to be damaging to the protein product. Furthermore, a number of genes within the RNO16 differential segment associated with metabolic syndrome components in human studies showed polymorphisms between SHR and BN-Lx (including Lpl, Nrg3, Pbx4, Cilp2, and Stab1). Our novel congenic rat model demonstrates that a limited genomic region on RNO16 in the SHR significantly affects many of the features of metabolic syndrome.

Brown Norway rats, a putative schizophrenia model, show increased electroencephalographic activity at rest and decreased event-related potential amplitude, power, and coherence in the auditory sensory gating paradigm.

In recent schizophrenia clinical research, electroencephalographic (EEG) oscillatory activities induced by a sensory stimulus or behavioral tasks have gained considerable interest as functional and pathophysiological biomarkers. The Brown Norway (BN) rat is a putative schizophrenia model that shows naturally low sensorimotor gating and deficits in cognitive performance, although other phenotypes have not been studied. The present study aimed to investigate the neurophysiological features of BN rats, particularly EEG/event-related potential (ERP). EEG activity was recorded at rest and during the auditory sensory gating paradigm under an awake, freely moving condition. Frequency and ERP analysis were performed along with time-frequency analysis of evoked power and intertrial coherence. Compared with Wistar-Kyoto rats, a well-documented control line, BN rats showed increased EEG power at rest, particularly in the theta and gamma ranges. In ERP analysis, BN rats showed reduced N40-P20 amplitude but normal sensory gating. The rats also showed reduced evoked power and intertrial coherence against auditory stimuli. These results suggest that BN rats show features of EEG/ERP measures clinically relevant to schizophrenia and may provide additional opportunities for translational research.

Distinct retinohypothalamic innervation patterns predict the developmental emergence of species-typical circadian phase preference in nocturnal Norway rats and diurnal nile grass rats.

How does the brain develop differently to support nocturnality in some mammals, but diurnality in others? To answer this question, one might look to the suprachiasmatic nucleus (SCN), which is entrained by light via the retinohypothalamic tract (RHT). However, because the SCN is more active during the day in all mammals studied thus far, it alone cannot determine circadian phase preference. In adult Norway rats (Rattus norvegicus), which are nocturnal, the RHT also projects to the ventral subparaventricular zone (vSPVZ), an adjacent region that expresses an in-phase pattern of SCN-vSPVZ neuronal activity. In contrast, in adult Nile grass rats (Arvicanthis niloticus), which are diurnal, an anti-phase pattern of SCN-vSPVZ neuronal activity is expressed. We hypothesized that these species differences result in part from a weak or absent RHT-to-vSPVZ projection in grass rats. Here, using a developmental comparative approach, we assessed species differences in behavior, hypothalamic activity, and RHT anatomy. We report that a robust retina-to-vSPVZ projection develops in Norway rats around the end of the second postnatal week when nocturnal wakefulness and the in-phase pattern of neuronal activity emerge. In grass rats, however, such a projection does not develop and the emergence of the anti-phase pattern during the second postnatal week is accompanied by increased diurnal wakefulness. When considered within the context of previously published reports on RHT projections in a variety of species, the current findings suggest that how and when the retina connects to the hypothalamus differentially shapes brain and behavior to produce animals that occupy opposing temporal niches.

The effects of buprenorphine on behaviour in the ACI and BN rat inbred strains.

Buprenorphine is a partial mu, kappa agonist that has been shown to influence spontaneous behaviour in animals. Previously, we have demonstrated significant differences in the analgesic response to buprenorphine between the August Copenhagen Irish (ACI)/SegHsd and the Brown Norway (BN)/RijHsd inbred rat strains. The purpose of this study was to determine whether these strains also differed in their behavioural response to buprenorphine in order to provide an additional parameter for the genetic analysis and localization of genes involved in this response. Male and female rats of both strains were used (n = 6/strain/sex) for this study. Each rat was subjected, respectively, to three treatment regimens at 15:00 h: (A) unchallenged; (B) intravenous saline; (C) intravenous buprenorphine (0.05 mg/kg) according to a crossover design. The relative duration (s/h) of locomotion, grooming, drinking and eating behaviour was subsequently determined from 15:30 to 07:00 h using the automatic registration system, Laboratory Animal Behaviour Registration and Analysis System(trade mark). Significant strain differences were observed in unchallenged behaviour between the ACI and the BN rats. ACI rats, but not BN rats, responded to buprenorphine treatment with decreased levels of locomotion, drinking and eating behaviour. The same treatment resulted in an increased grooming behaviour in both strains. Slight but significant sex differences were observed for locomotion and eating in the analysis of variance procedure, but did not reach the level of statistical significance in the multiple comparison procedure. The results of this study emphasize the possibility that strain-specific effects must be taken into account when using behavioural parameters for the assessment of the analgesic effects of buprenorphine in rats.

Age-related changes in cardiac structure and function in Fischer 344 x Brown Norway hybrid rats.

The effects of aging on cardiovascular function and cardiac structure were determined in a rat model recommended for gerontological studies. A cross-sectional analysis assessed cardiac changes in male Fischer 344 x Brown Norway F1 hybrid rats (FBN) from adulthood to the very aged (n = 6 per 12-, 18-, 21-, 24-, 27-, 30-, 33-, 36-, and 39-mo-old group). Rats underwent echocardiographic and hemodynamic analyses to determine standard values for left ventricular (LV) mass, LV wall thickness, LV chamber diameter, heart rate, LV fractional shortening, mitral inflow velocity, LV relaxation time, and aortic/LV pressures. Histological analyses were used to assess LV fibrotic infiltration and cardiomyocyte volume density over time. Aged rats had an increased LV mass-to-body weight ratio and deteriorated systolic function. LV systolic pressure declined with age. Histological analysis demonstrated a gradual increase in fibrosis and a decrease in cardiomyocyte volume density with age. We conclude that, although significant physiological and morphological changes occurred in heart function and structure between 12 and 39 mo of age, these changes did not likely contribute to mortality. We report reference values for cardiac function and structure in adult FBN male rats through very old age at 3-mo intervals.

Impaired myogenic autoregulation in kidneys of Brown Norway rats.

The Brown Norway (BN) rat is normotensive and has an extended lifespan but is extremely sensitive to hypertension-induced renal injury. Relative impairment of autoregulation has been implicated in the progression of renal failure whereas absence of myogenic autoregulation is associated with early renal failure. Therefore, we tested the hypothesis that there is conditional failure of renal autoregulation in BN rats. In isoflurane-anesthetized BN rats, the pressure-flow transfer function was normal when pressure fluctuated spontaneously. External forcing increased pressure fluctuation and exposed weakness of the myogenic component of autoregulation; the component mediated by tubuloglomerular feedback was less affected. In the presence of vasopressin to raise renal perfusion pressure, myogenic autoregulation was further impaired during forcing in BN rats but not in Wistar rats. Compensation by the myogenic system was rapidly restored on cessation of forcing, suggesting a functional limitation rather than a structural failure. Graded forcing in Wistar rats and in spontaneously hypertensive rats revealed that compensation due to the myogenic system was strong and independent of forcing amplitude. In contrast, graded forcing in BN rats showed that compensation was reduced when fluctuation of blood pressure was increased but that the reduction was independent of forcing amplitude. The results demonstrate conditional failure of myogenic autoregulation in BN rats. These acute studies provide a possible explanation for the observed sensitivity to hypertension-induced renal injury in BN rats.

Angiotensin I-converting enzyme genotype influences arterial response to injury in normotensive rats.

Two normotensive strains of rat, the Lou and Brown Norway (BN) strains, have contrasting levels of plasma angiotensin-converting enzyme (ACE). To investigate the degree of genetic determination of ACE expression, a polymorphic marker of the ACE gene was analyzed in inbred rats of the two strains. The two inbred strains were shown to bear different alleles for a polymorphic marker at the ACE gene. The segregation of the alleles of this marker and the plasma ACE levels were studied in a group of F2 rats issued from a cross between Lou and BN rats. The degree of genetic determination of plasma ACE activity was estimated to be 94% in the F2 cohort. The ACE locus accounts for 74% of total plasma ACE variance. ACE activity and mRNA expression in lungs were also genetically determined. The difference observed in ACE mRNA accumulation in the lungs between the two strains was due to a difference in the transcriptional rate of the ACE gene, as shown in nuclear run-on experiments. No differences were observed in arterial blood pressure of homozygous F2 progeny. In these animals, ACE genotype did not interfere with the pressor or the depressor responses to ACE-dependent vasoactive peptides. There was a significant effect of strain on constitutive or inducible membrane or soluble ACE activity in primary cultures of vascular cells. Neointima formation in the carotid artery 14 days after balloon injury was also influenced by the genotype in F2 homozygous progeny, whereas the medial area was not. These results demonstrate that there is a close relationship between the genetically determined ACE expression and the inducibility of the ACE gene. The degree of genetic determination of ACE expression in inbred rat strains offers a unique opportunity to study the interaction between genetic and environmental determinants of ACE expression and its involvement in response to experimental cardiovascular and renal injury.

Pronounced juvenile circadian core temperature rhythms exist in several strains of rats but not in rabbits.

Torpor-like circadian variations of core temperature are well documented for suckling-age Zucker rat pups. To determine (1) whether this juvenile circadian rhythm is as strongly expressed in other rat strains, and (2) whether a similar rhythm is expressed in rabbit pups, we recorded core temperature and metabolic rate of artificially reared pups. Wistar, Brown Norway, and Long Evans pups were studied for 30 h under moderate cold loads (ambient temperature = 28 degrees C) when 9-11 days old, i.e. at the age and ambient temperature for which the rhythm has been most thoroughly characterized in Zucker rats. Chinchilla bastard rabbit pups were studied under similar conditions when they were 3-8 days old, the youngest age at which the rhythm can be easily detected in rats. Rat pups of each strain showed clear circadian rhythms with sharp decreases of core temperature and metabolic rate in subjective morning. Core temperature amplitudes were in the order Wistar < Brown Norway < Zucker < Long Evans strain. In contrast, the rabbit pups maintained stable high levels of core temperature and metabolic rate throughout the day. A torpor-like decrease of core temperature in the morning is thus not a peculiarity of the Zucker rat strain but also occurs in other pigmented rat strains, whereas rabbit pups at a similar developmental stage do not show a circadian core temperature rhythm.

Role of cyclooxygenase and 5-lipoxygenase metabolites, platelet-activating factor and 5-hydroxytryptamine in allergen-induced airway responses in the brown Norway rat.

We determined the effects of selective inhibition of arachidonic acid metabolism via the cyclooxygenase and 5'-lipoxygenase pathways using flurbiprofen and BWA4C, respectively, of 5-hydroxytryptamine (5-HT) using methysergide and of platelet-activating factor (PAF) using WEB 2086 on the airway responses to ovalbumin (OA) aerosol in OA-sensitized Brown Norway rats. Twenty-one days after intraperitoneal injection of OA, rats were exposed to a 1% OA or saline aerosol. Only methysergide (10 mg/kg i.p.; 3 doses over 24 h) provided significant protection of the immediate response to OA. The increase in airway responsiveness to acetylcholine after OA exposure was not significantly altered by methysergide, flurbiprofen (10 mg/kg i.p.), BWA4C (50 mg/kg i.p.) and WEB 2086 (50 mg/kg i.p.) all given over 24 h prior to OA challenge. In addition, there was no effect on the increased recovery of eosinophils and lymphocytes in bronchoalveolar lavage fluid at 24 h. We conclude that 5-HT is an important mediator of the acute response to OA, but that 5-HT, lipoxygenase and cyclooxygenase products and PAF are unlikely to be involved in OA-induced airway hyperresponsiveness and inflammation in the Brown Norway rat.

HgCl2-induced glandular pathosis in the brown Norway rat.

Low doses of HgCl2 induce a genetically restricted autoimmune syndrome in Brown Norway (BN) rats (BN(Hg)). Part of the syndrome includes spontaneously developing mononuclear cell foci in salivary and lacrimal glands, morphologically similar to focal sialoadenitis in Sjögren's syndrome in man. In this study, we have shown that Hg-induced focal adenitis in BN rat is not female predominant and not solely dependent on Hg uptake in the glands. In BN(Hg), focal adenitis was found to develop in several different glands (parotid, submandibular, lacrimal, thyroid) as a specific manifestation of immunopathosis induced by Hg in the RT1n-haplotype-carrying BN. Such manifestations were absent in another strain, the Hg-resistant Lewis (RT1l). Also a potent contact sensitizer, DNFB, induced focal adenitis in isolated glands, provided that strong sensitization had occurred. Immunohistochemical characterization of focal adenitis in BN(Hg) with monoclonal antibodies showed that T cells and dendritic cells dominate the early infiltrates, whereas B cells were absent.

Non convulsive spike-wave discharges do not induce Fos in cerebro-cortical neurons.

Immunohistochemical localisation of Fos was used as a marker of neuronal activity to demonstrate neurons active during non-convulsive spike-wave epilepsy. Fos-positive neurons in cortex and several subcortical areas were counted. In undisturbed animals. Fos counts were not related to spike-wave in any region. With the electroencephalographic (EEG) recording procedure. Fos induction occurred in all regions, even after habituation. However, in central cortex, counts were found to be inversely related to spike-wave duration. This suggests that neuronal activity is not increased during spike-wave and that the central cortex in these animals is less responsive to arousal than in non-epileptic animals.

Platelet aggregation in spontaneous hypertension: genetic determination and correlation analysis.

OBJECTIVE: Hyper-responsive platelets are often found in essential hypertension. It has also been suggested that in hypertensive patients platelets may serve as an easily accessible indicator of abnormalities in contractile cell function. To test these suggestions, we analysed the relationship between platelet aggregation and genetic hypertension in the rat. METHODS: Linear regression analysis of mean values of recombinant inbred strains was used to evaluate the relationship between blood pressure and ADP-induced platelet aggregation. RESULTS: ADP-induced platelet aggregation in platelet-rich plasma in spontaneously hypertensive rats (SHR) was significantly decreased compared with in normotensive Brown Norway (BN) rats. However, in recombinant inbred strains, derived from (SHR x BN) F2 hybrids, correlation analysis revealed that platelet aggregation and blood pressure are independent traits. CONCLUSIONS: The present results suggest strongly that spontaneous hypertension and platelet hypo-aggregability in SHR were linked together by chance due to drift during selective inbreeding. The absence of a correlation between the two traits also indicates that alterations in platelet function in essential hypertension, often found in population-based studies, may have to be reaffirmed in genetically better-defined situations, e.g. by pedigree analysis.

The BN acute myelocytic leukemia (BNML) (a rat model for studying human acute myelocytic leukemia (AML)).

Even if animal models have many properties in common with the human disease, as is the case for the BNML and human AML, they have their limitations with respect to the extrapolation to the clinical situation. This also holds for the BNML; thus, conclusions should only be drawn with great caution. Nevertheless, the studies in the BNML model have added considerably to the understanding of various processes that occur during the development of leukemia, e.g., the interaction of leukemic cells and normal hemopoietic stem cells in relation to the microenvironment. The methodology developed in the BNML model allows the quantification of the relative effectiveness of any given treatment with regard to the antileukemic activity compared with the toxicity for normal host tissues. Furthermore, the cell kinetic studies performed in the BNML as a consequence of timed sequential chemotherapy has been helpful in designing an approach to take advantage of this phenomenon in the treatment of acute leukemia. The comparison of the various treatment modalities, employed for the conditioning prior to bone marrow transplantation, made it possible to determine the relative effectiveness of the various approaches. The fractionation of total body irradiation for conditioning purposes was supposed to have a negligible effect with regard to a reduced antileukemic effect. Detailed studies that were conducted in the BNML model did not confirm this hypothesis indicating that (hyper-)fraction of TBI results in a reduced antileukemic effect. The in vitro purging studies in the BNML aimed at the elimination of residual leukemic cells in autologous bone marrow transplantation contributed to the introduction of this method in clinical practice. However, extended studies in the BNML model also indicated that the contribution of the residual leukemia cell in the patient contributed to a much greater extend to the recurrence of leukemia then did the residual cells in the autologous marrow graft. A major contribution of the BNML was achieved in the study of the area of so-called "minimal residual disease" (MRD). A number of so-far unknown aspects of relapsing leukemia could be identified and studied. A new concept of discriminating locally relapsing leukemia and a delayed occurrence of generalized spreading of leukemia formed the basis for the explanation of the observed heterogeneity in the distribution of leukemic cells during the remission and the subsequent relapse phase. In conclusion, it is obvious that proper comparison of the human disease as well as the counterpart in the animal model requires a detailed knowledge of both.(ABSTRACT TRUNCATED AT 400 WORDS)

Strain differences in the sleep of rats.

Sleep was measured in two inbred rat strains (Lewis and Brown Norway) and their F1 hybrids to investigate patterns of inheritance and to provide a starting point for future studies of F2 and recombinant rats. Recordings from chronically implanted electrodes were quantified and scored by a computerized system; results were evaluated by an analysis of variance with pairwise comparisons by the Tukey HSD test. Brown Norway rats had the highest paradoxical sleep (PS) percentages; Lewis rats had the lowest. Hybrid rats had PS percentages intermediate between parent strains and significantly different from both. These results suggest codominance or multigenic transmission of PS amounts. There were no group differences of number of PS bouts; Brown Norway and hybrid rats had longer bouts than Lewis rats. Lewis and hybrid rats had similar amplitudes of the diurnal rhythm of PS, which were higher than those of Brown Norway rats; single gene transmission remains possible for diurnal rhythm amplitude. Thus, inheritance of PS percentage and rhythm amplitude appear independent. No group differences in PS latency were found.

Analysis of survival data on aging rat cohorts: pitfalls and some practical considerations.

Experimental aging research is very dependent on the determination of the survival characteristics of the animal species or strain under study. Such data are generally inferred from mortality curves of cohorts of animals that are set aside at an early age for aging studies. Rectangular survival curves and the presence of multiple pathological lesions are a prerequisite for aging studies so as to resemble the situation in man. From 1977 onwards, many rat cohorts have been formed in the Institute for Experimental Gerontology (IVEG) for the study of aging processes. Data from these have been analysed for a period of 5 years up to and including 1982. (Males and females of the WAG/Rij and BN/BiRij strains were used.) The 50% survival and the maximum survival of cohorts varied considerably, but showed no consistent trend over the years. The median (50%) survival between the cohorts differed by as much as 7.9-10.7 months for the strains and sexes studied. Maximum survival between the cohorts varied from 3.7 to 9.9 months. Median and maximal survival were greater for the females. Maximum survival and 50% survival correlated significantly, the relation between the two being approximately linear. The effect of removing animals from cohorts on the estimation of 50% survival was only minor, whereas maximum survival was clearly diminished by this procedure. The wide variation in survival characteristics, even between successive cohorts, cautions against too simple a measure of the animals survival in only one number for median or maximal survival in months. An indication of the variance of 50% survival and of maximum survival should therefore be included in scientific publications. Moreover, the 50% survival is the parameter of choice to define cohorts, not only because this can be most reliably estimated with good confidence limits, but also because this measure is the least sensitive to removing animals from the cohorts. As this will often be the case in many research institutions, it might be of practical importance to order old animals from different cohorts since this diminishes the chance of using an extremely short or long lived cohort. Finally, the analysis revealed that combining intact or incomplete cohorts into larger survival curves resulted in nearly identical graphs. An attempt was made to calculate the minimum cohort size which yields survival curves with constant 95% confidence limits.

Effect of prostaglandin E1 in brown Norway rats with mercury-induced autoimmune disease.

The effect of prostaglandin E1 on mercury-induced autoimmune disease in brown Norway rats has been investigated. Daily doses of 6 to 24 micrograms prolonged survival and significantly decreased proteinuria, deposition of immune reactants in the glomeruli, circulating anti-glomerular membrane antibody production, total serum IgE, and circulating immune complex level. A dose of 3 micrograms was also effective but to a lesser degree. These results show the efficiency of prostaglandin E1 in yet another autoimmune disease, show that the beneficial effect of prostaglandin E1 in this model is related to its immunosuppressive effects, and suggest that modification of prostaglandin-mediated suppression induced by HgCl2 might play a role in the pathogenesis of this autoimmune disease.

Prolonged activated partial thromboplastin time and deficiency of high molecular weight kininogen in brown Norway rat mutant (Katholiek strain).

Activated partial thromboplastin time (APTT) was examined in Brown Norway (B/N) Katholiek rat, which was previously reported as high molecular weight kininogen deficient. APTT of B/N Katholiek was prolonged to 35 sec in comparison with B/N Kitasato and SD rat, showing APTT of 22-24 sec. The mixture of B/N Katholiek plasma and B/N Kitasato plasma (1:1) showed normal APTT value. B/N Katholiek plasma corrected the abnormally prolonged human coagulation factor deficient plasmas, such as XI, XII and prekallikrein deficient plasmas, while it did not correct the APTT of HMW kininogen deficient, Fitzgerald plasma. Intravenous injection of bromelain, which was previously reported to produce prolonged hypotension through the activation of factor XII to release bradykinin, induced slight effect in Katholiek rat, while in Kitasato rat it showed prolonged hypotension in similar degree as SD rat. Contents of coagulation factors in B/N Katholiek thus measured as well as the values of prekallikrein and HMW kininogen previously reported were summarized and suggested that B/N Katholiek rat could be similar deficiency as Fitzgerald trait.