Inbred Rat Model Research Articles

Loss of the heterogeneous expression of flippase ATP11B leads to cerebral small vessel disease in a normotensive rat model

Cerebral small vessel disease (SVD) is the leading cause of vascular dementia, causes a quarter of strokes, and worsens stroke outcomes. The disease is characterised by patchy cerebral small vessel and white matter pathology, but the underlying mechanisms are poorly understood. This microvascular and tissue damage has been classically considered secondary to extrinsic factors, such as hypertension, but this fails to explain the patchy nature of the disease, the link to endothelial cell (EC) dysfunction even when hypertension is absent, and the increasing evidence of high heritability to SVD-related brain damage. We have previously shown the link between deletion of the phospholipase flippase Atp11b and EC dysfunction in an inbred hypertensive rat model with SVD-like pathology and a single nucleotide polymorphism (SNP) in ATP11B associated with human sporadic SVD. Here, we generated a novel normotensive transgenic rat model, where Atp11b is deleted, and show pathological, imaging and behavioural changes typical of those in human SVD, but that occur without hypertension. Atp11bKO rat brain and retinal small vessels show ECs with molecular and morphological changes of dysfunction, with myelin disruption in a patchy pattern around some but not all brain small vessels, similar to the human brain. We show that ATP11B/ATP11B is heterogeneously expressed in ECs in normal rat and human brain even in the same transverse section of the same blood vessel, suggesting variable effects of the loss of ATP11B on each vessel and an explanation for the patchy nature of the disease. This work highlights a link between inherent EC dysfunction and vulnerability to SVD white matter damage with a marked heterogeneity of ECs in vivo which modulates this response, occurring even in the absence of hypertension. These findings refocus our strategies for therapeutics away from antihypertensive (and vascular risk factor) control alone and towards ECs in the effort to provide alternative targets to prevent a major cause of stroke and dementia.

Addressing cancer invasion and cell motility with quantitative light microscopy

The incidence of death caused by cancer has been increasing worldwide. The growth of cancer cells is not the main problem. The majority of deaths are due to invasion and metastasis, where cancer cells actively spread from primary tumors. Our inbred rat model of spontaneous metastasis revealed dynamic phenotype changes in vitro correlating with the metastatic potential in vivo and led to a discovery of a metastasis suppressor, protein 4.1B, which affects their 2D motility on flat substrates. Subsequently, others confirmed 4.1B as metastasis suppressor using knock-out mice and patient data suggesting mechanism involving apoptosis. There is evidence that 2D motility may be differentially controlled to the 3D situation. Here we show that 4.1B affects cell motility in an invasion assay similarly to the 2D system, further supporting our original hypothesis that the role of 4.1B as metastasis suppressor is primarily mediated by its effect on motility. This is encouraging for the validity of the 2D analysis, and we propose Quantitative Phase Imaging with incoherent light source for rapid and accurate testing of cancer cell motility and growth to be of interest for personalized cancer treatment as illustrated in experiments measuring responses of human adenocarcinoma cells to selected chemotherapeutic drugs.

The reinforcement threshold and elasticity of demand for nicotine in an adolescent rat model of depression

BackgroundThe Food and Drug Administration (FDA) is considering setting a nicotine standard for tobacco products to reduce their addictiveness. Such a standard should account for the apparent greater vulnerability to nicotine addiction in some subpopulations, such as adolescents with depression. The present study examined whether the reinforcement threshold and elasticity of demand (i.e., reinforcing efficacy) for nicotine in a genetic inbred rat model of depression (Flinders Sensitive Line [FSL]) differs from an outbred control strain. MethodsAcquisition of nicotine self-administration (NSA) across a wide range of nicotine doses was measured in both FSL and Sprague-Dawley (SD) control adolescent rats. At the highest dose, elasticity of demand was also measured. Nicotine pharmacokinetics was examined to determine whether it might modulate NSA, as it does smoking in humans. ResultsFSL rats acquired self-administration quicker and showed more inelastic demand (greater reinforcing efficacy) than SDs at the highest unit dose. However, there was no strain difference in the reinforcement threshold of nicotine. FSL rats exhibited faster nicotine clearance, larger volume of distribution, and lower plasma and brain nicotine concentrations. However, these differences were not consistently related to strain differences in NSA measures. ConclusionThese findings are consistent with studies showing greater dependence and reinforcing efficacy of cigarettes in smokers with depression and those with relatively fast nicotine metabolism. However, these findings also suggest that a nicotine standard to reduce initiation of tobacco use should be similarly effective in both the general adolescent population and those with depression.

Pulling the Hood off Genetic Susceptibility to Hypertensive Renal Disease

Pulling the Hood off Genetic Susceptibility to Hypertensive Renal Disease

Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage.

Unraveling the genetic susceptibility of complex diseases such as chronic kidney disease remains challenging. Here, we used inbred rat models of kidney damage associated with elevated blood pressure for the comprehensive analysis of a major albuminuria susceptibility locus detected in these models. We characterized its genomic architecture by congenic substitution mapping, targeted next-generation sequencing, and compartment-specific RNA sequencing analysis in isolated glomeruli. This led to prioritization of transmembrane protein Tmem63c as a novel potential target. Tmem63c is differentially expressed in glomeruli of allele-specific rat models during onset of albuminuria. Patients with focal segmental glomerulosclerosis exhibited specific TMEM63C loss in podocytes. Functional analysis in zebrafish revealed a role for tmem63c in mediating the glomerular filtration barrier function. Our data demonstrate that integrative analysis of the genomic architecture of a complex trait locus is a powerful tool for identification of new targets such as Tmem63c for further translational investigation.

Inbred Rats as a Model to Study Persistent Renal Leptospirosis and Associated Cellular Immune Responsiveness.

Pathogenic species of Leptospira cause leptospirosis, a bacterial zoonotic disease with a global distribution affecting over one million people annually. Rats are regarded as one of the most significant reservoir hosts of infection for human disease, and in the absence of clinical signs of infection, excrete large numbers of organisms in their urine. A unique biological equilibrium exists between pathogenic leptospires and reservoir hosts of infection, but surprisingly, little is known concerning the host's cellular immune response that facilitates persistent renal colonization. To address this deficiency, we established and applied an immunocompetent inbred rat model of persistent renal colonization; leptospires were detected in urine of experimentally infected rats by 3 weeks post-infection and remained positive until 8 weeks post-infection. However, there was little, if any, evidence of inflammation in colonized renal tubules. At 8 weeks post-infection, a robust antibody response was detected against lipopolysaccharide and protein outer membrane (OM) components. Purified B and T cells derived from the spleen of infected and non-infected rats proliferated in response to stimulation with 0.5 μg of OM fractions of Leptospira, including CD4+ T cells, which comprised 40% of proliferating cells, compared to 25% in non-infected controls. However, analysis of gene expression did not determine which immunoregulatory pathways were activated. Lymphocytes purified from the lymph node draining the site of colonization, the renal lymph node, also showed an increase in percentage of proliferating B and T cells. However, in contrast to a phenotype of 40% CD4+ T cells in the spleen, the phenotype of proliferating T cells in the renal lymph node comprised 65% CD4+ T cells. These results confirm that the renal lymph node, the local lymphoid organ, is a dominant site containing Leptospira reactive CD4+ T cells and highlight the need to consider the local, vs. systemic, immune responses during renal colonization infection. The use of inbred immunocompetent rats provides a novel tool to further elucidate those pathophysiological pathways that facilitate the unique biological equilibrium observed in reservoir hosts of leptospirosis.

Integrative genomic, phenomic and transcriptomic study of metabolic syndrome in recombinant inbred rat model set

Integrative genomic, phenomic and transcriptomic study of metabolic syndrome in recombinant inbred rat model set

Identification of a nutrient-sensing transcriptional network in monocytes by using inbred rat models on a cafeteria diet.

ABSTRACTObesity has reached pandemic levels worldwide. The current models of diet-induced obesity in rodents use predominantly high-fat based diets that do not take into account the consumption of variety of highly palatable, energy-dense foods that are prevalent in Western society. We and others have shown that the cafeteria (CAF) diet is a robust and reproducible model of human metabolic syndrome with tissue inflammation in the rat. We have previously shown that inbred rat strains such as Wistar Kyoto (WKY) and Lewis (LEW) show different susceptibilities to CAF diets with distinct metabolic and morphometric profiles. Here, we show a difference in plasma MCP-1 levels and investigate the effect of the CAF diet on peripheral blood monocyte transcriptome, as powerful stress-sensing immune cells, in WKY and LEW rats. We found that 75.5% of the differentially expressed transcripts under the CAF diet were upregulated in WKY rats and were functionally related to the activation of the immune response. Using a gene co-expression network constructed from the genes differentially expressed between CAF diet-fed LEW and WKY rats, we identified acyl-CoA synthetase short-chain family member 2 (Acss2) as a hub gene for a nutrient-sensing cluster of transcripts in monocytes. The Acss2 genomic region is significantly enriched for previously established metabolism quantitative trait loci in the rat. Notably, monocyte expression levels of Acss2 significantly correlated with plasma glucose, triglyceride, leptin and non-esterified fatty acid (NEFA) levels as well as morphometric measurements such as body weight and the total fat following feeding with the CAF diet in the rat. These results show the importance of the genetic background in nutritional genomics and identify inbred rat strains as potential models for CAF-diet-induced obesity.

The complete mitochondrial genome sequence of an endometrial cancer inbred Donryu rat model

The Donryu rat strain is a commonly used model for endometrial cancer disease study. We sequenced this rat strain mitochondrial genome for the first time (GenBank Accession No. KM114605). Its mitogenome was 16,307 bp and coding 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes. A total of 96 SNPs were examined when compared to reference BN sequence.

Autism-related behavioral phenotypes in an inbred rat substrain

Behavioral and genetic differences among Wistar–Kyoto (WKY) rats from different vendors and different breeders have long been observed, but generally overlooked. In our prior work, we found that two closely related WKY substrains, the WKY/NCrl and WKY/NHsd rats, differ in a small percentage of their genome which appeared to be highly enriched for autism risk genes. Although both substrains have been used widely in studies of hypertension, attention deficit/hyperactivity disorder (ADHD) and depression, they have not been tested for any autism-related behavioral phenotypes. Furthermore, these two substrains have often been used interchangeably in previous studies; no study has systematically examined the phenotypic differences that could be attributed by their small yet potentially meaningful genetic differences. In this paper we compared these two substrains on a battery of neurobehavioral tests. Although two substrains were similar in locomotor activity, WKY/NCrl rats were significantly different from WKY/NHsd rats in the elevated plus maze test, as well as measures of social interaction and ultrasonic vocalization. These strains were also compared with Sprague Dawley (SD) rats, a common outbred strain, and spontaneous hypertensive rats (SHR), an inbred rat model for ADHD and hypertension, which were derived from the same ancestor strain as the WKY strains. Our behavioral findings suggest that WKY/NCrl rats may be useful as a model autism spectrum disorders due to their lower social interest, lower ultrasonic vocalization and higher anxiety levels when WKY/NHsd rats are used as the control strain. Given the small genetic difference between the two inbred substrains, future studies to identify the exact gene and sequence variants that differ between the two may be useful for identifying the genetic mechanisms underlying these behaviors.

Modularization and epistatic hierarchy determine homeostatic actions of multiple blood pressure quantitative trait loci

Hypertension, the most frequently diagnosed clinical condition world-wide, predisposes individuals to morbidity and mortality, yet its underlying pathological etiologies are poorly understood. So far, a large number of quantitative trait loci (QTLs) have been identified in both humans and animal models, but how they function together in determining overall blood pressure (BP) in physiological settings is unknown. Here, we systematically and comprehensively performed pair-wise comparisons of individual QTLs to create a global picture of their functionality in an inbred rat model. Rather than each of numerous QTLs contributing to infinitesimal BP increments, a modularized pattern arises: two epistatic 'blocks' constitute basic functional 'units' for nearly all QTLs, designated as epistatic module 1 (EM1) and EM2. This modularization dictates the magnitude and scope of BP effects. Any EM1 member can contribute to BP additively to that of EM2, but not to those of the same module. Members of each EM display epistatic hierarchy, which seems to reflect a related functional pathway. Rat homologues of 11 human BP QTLs belong to either EM1 or EM2. Unique insights emerge into the novel genetic mechanism and hierarchy determining BP in the Dahl salt-sensitive SS/Jr (DSS) rat model that implicate a portion of human QTLs. Elucidating the pathways underlying EM1 and EM2 may reveal the genetic regulation of BP.

Mapping genetic determinants of kidney damage in rat models

During the last two decades, significant progress in our understanding of the development of kidney diseases has been achieved by unravelling the mechanisms underlying rare familial forms of human kidney diseases. Due to the genetic heterogeneity in human populations and the complex multifactorial pathogenesis of the disease phenotypes, the dissection of the genetic basis of common chronic kidney diseases (CKD) remains a difficult task. In this regard, several inbred rat models provide valuable complementary tools to uncover the genetic basis of complex renal disease phenotypes that are related to common forms of CKD. In this review, data obtained in nine experimental rat models, including the Buffalo (BUF), Dahl salt-sensitive (SS), Fawn-hooded hypertensive (FHH), Goto-Kakizaki (GK), Lyon hypertensive (LH), Munich Wistar Frömter (MWF), Sabra hypertension-prone (SBH), spontaneously hypertensive rat (SHR) and stroke-prone spontaneously hypertensive rat (SHRSP) inbred strains, that contributed to the genetic dissection of renal disease phenotypes are presented. In this panel of inbred strains, a large number of quantitative trait loci (QTL) linked to albuminuria/proteinuria and other functional or structural kidney abnormalities could be identified by QTL mapping analysis and follow-up studies including consomic and congenic rat lines. The comprehensive exploitation of the genotype-renal phenotype associations that are inherited in this panel of rat strains is suitable for making a significant contribution to the development of an integrated approach to the systems genetics of common CKD.

Linkage study of embryopathy—Polygenic inheritance of diabetes-induced skeletal malformations in the rat

We developed an inbred rat model of diabetic embryopathy, in which the offspring displays skeletal malformations (agnathia or micrognathia) when the mother is diabetic, and no malformations when she is not diabetic. Our aim was to find genes controlling the embryonic maldevelopment in a diabetic environment.We contrasted the fetal outcome in inbred Sprague-Dawley L rats (20% skeletal malformations in diabetic pregnancy) with that of inbred Wistar Furth rats (denotedW, no skeletal malformations in diabetic pregnancy). We used offspring from the backcross F1×L to probe for the genetic basis for malformation of the mandible in diabetic pregnancy. A set of 186 fetuses (93 affected, 93 unaffected) was subjected to a whole genome scan with 160 micro satellites. Analysis of genotype distribution indicated 7 loci on chromosome 4, 10 (3 loci), 14, 18, and 19 in the teratogenic process (and 14 other loci on 12 chromosomes with less strong association to the malformations), several of which contained genes implicated in other experimental studies of diabetic embryopathy. These candidate genes will be scrutinized in further experimentation.We conclude that the genetic involvement in rodent diabetic embryopathy is polygenic and predisposing for congenital malformations.

Strain differences in self-administration of methylphenidate and sucrose pellets in a rat model of attention-deficit hyperactivity disorder

Despite its abuse potential, methylphenidate (MPH) is widely prescribed for treatment of attention-deficit hyperactivity disorder (ADHD). The purpose of the present study was to examine MPH self-administration in a rat model of ADHD. Experiment 1 examined the acquisition of MPH self-administration and assessed the MPH dose-effect curve in spontaneously hypertensive rats (SHR), an inbred rat model of ADHD, Wistar Kyotos (WKY), the progenitor strain for SHR, and Sprague-Dawley (SD), an outbred control strain. Experiment 2 replicated Experiment 1, but replaced MPH infusions with sucrose pellets. Initial acquisition of MPH self-administration was greater in SHR and SD than WKY. However, with extended training using an incrementing fixed ratio schedule SHR and WKY did not differ in responding for MPH using the training dose (0.3 mg/kg/infusion) or other unit doses, except that SHR showed greater responding than WKY at 0.1 mg/kg/infusion. SHR also showed greater acquisition and maintenance of sucrose-reinforced responding compared with both WKY and SD. Greater initial acquisition of MPH self-administration in SHR than WKY may not be due to a strain-specific difference in sensitivity to the reinforcing effect of MPH.

The characteristics of rabbit and rat mesenchymal stromal cell growth and attachment to mesh used in hernia repair

The goal of this project is to create an optimal environment for wound healing. Preliminary results from a rabbit model suggest that the addition of mesenchymal stromal cells (MSC) to a repaired incision improves the biomechanical properties of the wound. To increase sample size and reproducibility, these studies will be repeated in an inbred rat model. A review of the literature shows a wide variety of culture conditions used for rat MSCs. This study compares proliferation of rabbit and rat MSCs when grown in ten different growth medias. The Cell Titer 96 ® Cell Proliferation Assay and direct cell counts were used to measure the proliferation of MSC's. Rabbit and rat MSC growth was optimal in alpha MEM + 20% FCS or Hams F12 + 10% FCS. To prepare for the extension of these studies into the rat model, four mesh materials commonly used in hernia repair were tested to determine the best vehicle for the addition of MSCs. MSCs were incubated with the mesh for 30 min at 37oC. Non‐adherent cells were removed and the remaining cell number determined by incubation with Cell Titer 96 ® solution and comparison to an MSC standard curve. Optimal adherence was seen with Bard ® CollaMend (38% of added cells). Future studies will use Bard ® CollaMend as a vehicle when adding MSCs to a repaired abdominal incision. Supported by St. Elizabeth Health Center and Youngstown State University.

Effect of glycosidation of isoflavones on their bioavailability and pharmacokinetics in aged male rats

There are limited reports on the bioavailability and pharmacokinetics of isoflavones in elderly humans and aged animals. The present study was conducted to assess the effect of glycosidation of isoflavones on their bioavailability and pharmacokinetics in aged (20 month old) male Fischer-344 (F-344) rats. The F-344 rat, developed by the National Institute on Aging, is an inbred rat model that is commonly used for aging studies and resembles many features of aging humans. Three sources of isoflavones; Novasoy (a commercial supplement), a mixture of synthetic aglycons (daidzein, genistein and glycitein), and a mixture of synthetic glucosides (daidzin, genistin, and glycitin) were tested. Following administration, blood samples were collected at different times (0-48 h post-oral gavage and 0-8 h post-IV dosing). Plasma isoflavones and 7-hydroxy-3-(4'-hydroxyphenyl)-chroman (a metabolite of daidzein) were measured by LC/MS. The extent of absorption was determined by comparing the area under the curve (AUC) of the plasma-concentration time curve after intravenous (IV) administration with that following oral administration. The extent of bioavailability was then calculated as: %bioabailability = (AUC(or)/AUC(IV))x(Dose(IV)/Dose(or))x100. Bioavailabilities for genistein were significantly (p = 0.013) higher for the aglycon (35 +/- 9%) compared with the glucoside forms (11 +/- 3%). In contrast, the bioavailabilities for glycitein were significantly (p = 0.011) higher in Novasoy (27 +/- 13%) and the glucoside form (21 +/- 10%) compared with the aglycon (8 +/- 3%). No significant differences in the bioavailability of daidzein were observed in aged rats dosed with aglycon, glucoside or Novasoy. However, aged rats were able to produce equol as early as 8 h post-dosing. In summary, the source of isoflavones had significant effects on genistein and glycitein bioavailability in aged male rats.

Dietary n‐3 fatty acids attenuate cardiac allograft vasculopathy via activating peroxisome proliferator‐activated receptor‐γ

Recent in vitro data suggested that n-3 fatty acids could inhibit the activation of PPAR gamma. This study was designed to test the hypothesis that fish oil ameliorates CAV development via activating PPAR gamma in an inbred rat model of heart transplantation. Animals were divided into four groups: isograft, control (CsA + vehicle), LFO-treated group (CsA + 0.3% v/w fish oil), and HFO-treated group (CsA + 0.6% v/w fish oil). CsA was administered at 1.5 mg/kg/day for two wk postoperatively. Recipients were treated with fish oil or vehicle daily for eight wk. The histopathological and immunohistochemical examination, activity of NF-kappaB and PPAR gamma, intragraft chemokine levels, and chemokine receptor expression were analyzed. Both LFO and HFO significantly decreased the CAV score, inhibited recruitment of T lymphocytes and macrophages, elevated the activity of PPAR gamma, inhibited the activity of NF-kappaB, reduced levels of intragraft MCP-1 and IP-10 as well as downregulated expression of chemokine receptors CCR2. CXCR3 expression was not affected. Our results demonstrated that fish oil might attenuate CAV development, possibly through activating PPAR gamma and subsequently inhibiting the NF-kappaB activation, the chemokines secretion, as well as the CCR2 expression.

Simvastatin attenuates cardiac isograft ischemia-reperfusion injury by down-regulating CC chemokine receptor-2 expression

Accumulating evidence reveals that statins possess direct anti-inflammatory properties through inhibition of proinflammatory cytokine and chemokine secretion in addition to their antioxidant effects, which may contribute to amelioration of ischemia-reperfusion injury. This study tested the hypothesis that perioperative treatment of simvastatin suppresses the cardiac isograft ischemia-reperfusion injury by down-regulation of CC chemokine receptor-2 expression in an inbred rat model of cardiac transplantation. Donor hearts from Lewis rats were heterotopically transplanted to Lewis rat recipients. Recipients were orally treated with simvastatin (1 mg/kg) or vehicle every morning 3 days before the surgery until the harvest day. Rats were killed at 6 hours and at 1, 3, and 7 days after transplantation. Injury was assessed by infarct size measurement, histologic and immunohistochemical examination, and intragraft myeloperoxidase activity assay. Monocyte chemoattractant protein-1 levels in serum and graft were analyzed by enzyme-linked immunosorbent assay, and intragraft CC chemokine receptor-2 expression was measured by quantitative real-time polymerase chain reaction. The infarct size and macrophage infiltration were all significantly reduced in the simvastatin-treated group compared with those of the control group at 1 day after transplantation. Neutrophil accumulation was significantly suppressed until 3 days after transplantation, whereas myeloperoxidase activity had been significantly diminished at 1 day after transplantation. Both monocyte chemoattractant protein-1 concentrations in serum and graft were remarkably decreased at 6 hours after transplantation. Intragraft CC chemokine receptor-2 expression was also down-regulated at 1 day and 3 days after transplantation. Perioperative treatment of simvastatin could suppress the isograft ischemia-reperfusion injury through retarding intragraft monocyte chemoattractant protein-1 accumulation and CC chemokine receptor-2 expression.

Inhibition of Chemokine Receptor CCR2 and CCR5 Expression Contributes to Simvastatin-induced Attenuation of Cardiac Allograft Vasculopathy

Accumulating evidence reveals that statins possess pleiotropic properties beyond cholesterol reduction, which may contribute to the attenuation of cardiac allograft vasculopathy (CAV). Recent in vitro data suggest that statins could down-regulate chemokine receptors. This study was designed to test the hypothesis that simvastatin ameliorates CAV development via the inhibition of chemokine receptor expression in an inbred rat model of cardiac transplantation. Animals were divided into four groups: isograft; control (cyclosporine [CsA] + vehicle); low-dose simvastatin (LSIM; CsA + 5 mg/kg simvastatin); and high-dose simvastatin (HSIM; CsA + 10 mg/kg simvastatin). Donor hearts from Fisher 344 rats were transplanted heterotopically into Lewis rat recipients. CsA was administered at 1.5 mg/kg/day for 2 weeks post-operatively. In addition, recipients were treated daily with simvastatin or vehicle for 8 weeks. Donor hearts were harvested for histopathologic and immunohistochemical examination. Intragraft concentration of chemokines and chemokine receptor expression were analyzed using enzyme-linked immunosorbent assay and quantitative real-time polymerase chain reaction, respectively. Both low and high doses of simvastatin significantly decreased the CAV score; inhibited recruitment of T lymphocytes and macrophages; reduced levels of intragraft MCP-1 (monocyte chemoattractant protein-1), RANTES (regulated on activation, normal T-cell expressed and secreted) protein and IP-10 (interferon-inducible protein-10); and down-regulated expression of chemokine receptors CCR2 and CCR5. CXCR3 expression was not affected by simvastatin treatment. Our results demonstrate that simvastatin may attenuate CAV development, possibly through retarding intragraft chemokine accumulation and chemokine receptor expression.

Positional Cloning of Quantitative Trait Loci for Blood Pressure: How Close Are We?

Blood pressure (BP) in any human population exhibits as a continuous variable that fits a bell-shaped curve. Hypertensive individuals are those whose BP is maintained at one extreme of the curve and above a defined cutoff. Despite progress made in identifying the mechanisms underlying certain rare monogenic forms of hypertension,1,2 the etiology and pathogenesis of essential hypertension remain poorly understood. Because existing human populations are genetically heterogeneous, and because environmental factors impacting on the pathogenesis of hypertension cannot be controlled in a given population, it is difficult to identify the molecular mechanisms that transduce the sequela of essential hypertension via direct human studies.3 To alleviate the drawbacks of human investigations, animal models, especially inbred rodents, have been developed and experimentally manipulated to identify quantitative trait loci (QTLs) for BP, because major confounding environmental factors, such as diet and genetic background, can be systematically controlled. Once identified in animal models, the molecular basis may be translated into physiological understandings of essential hypertension in humans. It is with this expectation that efforts have been launched to identify the molecular basis of BP QTLs in animal models. Because the identification of individual QTLs is primarily based on their chromosome locations unbiased by, or unrestricted to, their physiological roles, positional cloning is believed to be the most efficient strategy. Before we embark on discussions regarding QTL discovery, a definition is in order. Semantic arguments abound as to exactly what a QTL, that is, a locus,4 entails. Is it 1 gene or a collection of genes? As genetic mapping progresses from a large chromosome segment to an interval of submegabase, several regions initially thought to contain 1 BP QTL5 appear to harbor >1 in each of them,6–10 whereas several other regions turned out to harbor 1 QTL as expected.11,12 …