Background The recent development of a large panel of genome-wide single nucleotide polymorphisms (SNPs) provides the opportunity to examine genetic relationships between distinct SHR lines that share hypertension, but differ in their susceptibility to hypertensive end-organ disease. Methods and Results We compared genotypes at nearly 10,000 SNPs obtained for the hypertension end-organ injury-susceptible SHR-A3 (SHRSP, SHR-stroke prone) line and the injury-resistant SHR-B2 line. This revealed that that the two lines were genetically identical by descent (IBD) across 86.6% of the genome. Areas of the genome that were not IBD were distributed across 19 of the 20 autosomes and the X chromosome. A block structure of non-IBD comprising a total of 121 haplotype blocks was formed by clustering of SNPs inherited from different ancestors. To test the null hypothesis that distinct SHR lines share a common set of hypertension susceptibility alleles we compared blood pressure in adult SHR animals from both lines and their F1 and F2 progeny using telemetry. In 16–18wk old animals fed a normal diet, systolic blood pressure (SBP, mm Hg) in SHR-A3 was 205.7 ± 3.86 (mean ± SEM, n = 26), while in similar SHR-B2 animals SBP was 186.7 ± 2.53 (n = 20). In F1 and F2 animals, SBP was 188.2 ± 4.23, (n = 19) and 185.6 ± 1.1 (n = 211) respectively (p<10−6, ANOVA). In order to identify non-IBD haplotype blocks contributing to blood pressure differences between these SHR lines we developed a high throughput SNP genotyping system to genotype SNPs marking non-IBD blocks. We mapped a single non-IBD block on chromosome 17 extending over less than 10Mb at which SHR-A3 alleles significantly elevate blood pressure compared with SHR-B2. Conclusions Thus hypertension in SHR-A3 and -B2 appears to arise from an overlapping set of susceptibility alleles, with SHR-A3 possessing an additional hypertension locus that contributes to further increase blood pressure.

The decapeptide GnRH that regulates reproduction in all vertebrates is stored in, and secreted from, large dense-core secretory vesicles in nerve terminals in the median eminence. GnRH is released from these terminals with biological rhythms that are critical for the maintenance of normal reproduction. During reproductive aging in female rats, there is a loss of GnRH pulses and a diminution of the GnRH surge. However, information about the specific role of GnRH nerve terminals is lacking, particularly in the context of aging. We sought to gain novel ultrastructural information about GnRH neuroterminals by performing three-dimensional (3D) reconstructions of GnRH neuroterminals and their surrounding microenvironment in the median eminence of young (4-5 month) and old (22-24 month) ovariectomized Sprague-Dawley female rats. Median eminence tissues were freeze-plunge embedded, and serial ultrathin sections were collected on slot grids for immunogold labeling of GnRH immunoreactivity. Sequential images were used to create 3D models of GnRH terminals. These reconstructions provided novel perspectives into the morphological properties of GnRH terminals, and their neural and glial environment. We also noted that the cytoarchitectural features of the median eminence became disorganized with aging. Quantitative measures showed a significant decrease in the apposition between GnRH terminal membranes and glial cells. Our data suggest reproductive aging in rats is characterized by structural organizational changes to the GnRH terminal microenvironment in the median eminence.

Background The spontaneously hypertensive rat (SHR) strain exists in lines that contrast strongly in susceptibility to renal injury in hypertension. These inbred lines share common ancestry and only 13% of their genomes arise from different ancestors. Methods and Results We used next gen sequencing to detect natural allelic variation in 5 genes of the immunoreceptor signaling pathway (IgH, Dok3, Src, Syk and JunD) that arise from different ancestors in the injury-prone SHR-A3 and the resistant SHR-B2 lines. We created an intercross between these lines and in the F2 progeny we observed that the inheritance of haplotype blocks containing the SHR-A3 alleles of these 5 genes correlated with increased albuminuria and histological measures of renal injury. To test whether accumulated genetic variation in this pathway may create a therapeutic target in hypertensive renal injury, rats of both lines were treated with the immunosuppressant mycophenolate mofetil (MMF). MMF reduced proteinuria (albumin to creatinine ratio, uACR) from 6.6 to 1.2 mg/mg (p<0.001) in SHR-A3. Glomerular injury scores were reduced in MMF treated SHR-A3 from 1.6 to 1.4 (p<0.002). Tubulo-interstitial injury was reduced in MMF-treated SHR-A3 from 2.62 to 2.0 (p=0.001). MMF treatment also reduced renal fibrosis in SHR-A3, (3.9 vs. 2.0, p<0.001). Conclusions Polygenic susceptibility to renal injury in hypertension arises in association with genetic variation in genes that participate in immune responses and is dramatically improved by reduction of immune system activity.

The interaction between IgG and Fc-␥ receptors in glomeruli contributes to the development of several types of proteinuric glomerular disease, but the involvement of immunological mechanisms in hypertensive renal injury is incompletely understood. Here, we investigated serum IgG levels in SHR-A3 rats, which develop hypertensive injury, and compared them with the injury-resistant SHR-B2 line. At 18 weeks old, SHR-A3 rats had serum total IgG levels nearly twice those of SHR-B2 rats, although subclass IgG2b was undetectable in SHR-A3 rats compared with mean levels (Ϯ SEM) of 80.7 Ϯ 12.8 mg/dl (18 weeks) and 116.6 Ϯ 19.0 mg/dl (30 weeks) in SHR-B2 rats. In addition, these two strains had significantly different serum levels of IgG1, IgG2a, and IgG2c; differences persisted at 30 weeks for all subclasses except IgG2a. Genetic mapping revealed that a locus on chromosome 6 linked to IgG subclass levels that affected IgG1, IgG2b, and IgG2c but not IgG2a. The mapped haplotype block contains IgH, suggesting regulation of three of four serum IgG subclass levels in cis. Resequencing revealed variation in the sequence of the Fc portion of the IgG heavy chain, which predicts important functional changes. To examine whether there is any relationship between this haplotype block and susceptibility to renal injury, we examined the effect of SHR-A3 and SHR-B2 alleles at this block on albumin excretion in an F2 intercross. Albuminuria doubled with inheritance of SHR-A3 alleles. In summary, allelic variation in IgH or nearby genes may modulate the susceptibility to hypertensive renal injury in SHR-A3 rats.

Rapid assessment of adrenal function is critical following transsphenoidal surgery (TSS) for Cushing's disease (CD) in order to determine surgical efficacy. We hypothesize that there may be a role for ACTH measurement as a rapid indicator of adrenal function. Following surgery for CD, glucocorticoids were withheld and paired plasma ACTH and serum cortisol levels were measured every 6 h. Post-operative hypocortisolemia was defined as serum cortisol <2 mcg/dl or a serum cortisol <5 mcg/dl with the onset of symptoms of adrenal insufficiency within 72 h. We studied 12 subjects, all female, mean age 44.6 years (range 25-55), including 13 surgeries: nine subjects attained hypocortisolemia. Plasma ACTH levels decreased more in subjects with hypocortisolemia (0.9 pg/ml/hr, P = 0.0028) versus those with persistent disease (0 0.2 pg/ml/hr, P = 0.26) within the first 48 h after surgery. In contrast to subjects with persistent disease, all subjects with hypocortisolemia achieved a plasma ACTH <20 pg/ml by 19 h (range 1-19 h). Four of the nine subjects with hypocortisolemia achieved plasma ACTH <20 pg/ml by 13 h and the remaining five subjects by 19 h. Hypocortisolemia occurred between 3-36 h following achievement of a plasma ACTH <20 pg/ml. In CD, a reduction in postoperative plasma ACTH levels differentiates subjects with surgical remission versus subjects with persistent disease. The utility of plasma ACTH measurements in the postoperative management of CD remains to be determined.