Struvite Urolithiasis in Long-Evans Rats.

Struvite urinary calculi, which are composed of magnesium, ammonium, and phosphate, can cause complications including sepsis and renal failure. Struvite calculi were identified within the urinary bladder and renal pelvis of 2 Long-Evans rats that died within days after arrival from a commercial vendor. The remaining rats in the shipment were screened by physical examination, radiography, and ultrasonography, revealing an additional 2 animals that were clinically affected. These rats were euthanized, necropsied, and yielded similar findings to those from the first 2 rats. In addition, urine samples had an alkaline pH and contained numerous bacteria (predominantly Proteus mirabilis), leukocytes, and crystals. All calculi were composed completely of struvite. Another 7 rats in the shipment had alkaline urine with the presence of blood cells; 6 of these rats also had abundant struvite crystals, and P. mirabilis was cultured from the urine of 3 rats. Further investigation by the vendor identified 2 of 100 rats with struvite calculi from the same colony. Although no specific cause could be implicated, the fact that all the affected rats came from the same breeding area suggests a genetic or environmental triggering event; a contribution due to diet cannot be ruled out. Our findings suggest that the affected rats had metabolic disturbances coupled with bacterial infection that predisposed them to develop struvite calculi. During sudden increases of struvite urinary calculi cases in rats, urine cultures followed by appropriate surgical intervention and antibiotic therapy is warranted. Additional factors, including diet, merit attention as well.

Administration of luteinizing hormone releasing hormone agonist for synchronization of estrus and generation of pseudopregnancy for embryo transfer in rats.

In the past decade, the use of genetically engineered rats has increased exponentially; therefore, the ability to perform embryo transfer (ET) in rats to rederive, reanimate, or create mutant rat lines is increasingly important. However, the successful generation of pseudopregnant female rats for ET represents a limiting factor. We here evaluated the subcutaneous administration of 40 µg luteinizing hormone releasing hormone agonist (LHRHa) for estrus synchronization during the development and implementation of a rat ET program. Our first experiment assessed endogenous estrus cycling patterns by examining vaginal cytology without administration of LHRHa in 5-wk-old peripubertal Sprague-Dawley female rats. These rats then received LHRHa at approximately 7 wk of age; 57% of the rats were synchronized in proestrus or estrus as assessed by vaginal cytology 96 h later. In a second experiment, 8-wk-old virgin, unmanipulated Sprague-Dawley female rats received LHRHa; 55% were synchronized in proestrus or estrus 96 h later. Copulatory plugs were confirmed in 28% and 82% of the rats that had been synchronized in the first and second experiments, respectively, and mated with vasectomized male rats. Embryo transfer surgery was performed, and live pups were born from both fresh and cryopreserved transgenic rat embryos. Our results indicate that subcutaneous administration of 40 µg LHRHa followed by examination of vaginal cytology 96 h later is an effective technique to generate multiple pseudopregnant recipient rats for use in an ET program.

Restriction of endogenous T cell antigen receptor beta rearrangements to Vbeta14 through selective recombination signal sequence modifications.

T cell antigen receptor (TCR)beta V(D)J variable region exon assembly is ordered, with Dbeta to Jbeta rearrangements occurring before joining of Vbetas to a DJbeta complex. Germ-line V(D)J segments are flanked by recombination signal (RS) sequences, which consist of heptamers and nonamers separated by a spacer of 12 (12-RS) or 23 (23-RS) bp. V(D)J recombination is restricted by the 12/23 rule; joining occurs only between gene segments flanked by 12-RSs and 23-RSs. Vbeta segments have 23-RSs and Jbeta segments 12-RSs, which based on the 12/23 rule should allow direct joining. However, Vbeta segments rearrange only to DJbeta complexes and not Jbeta segments, because of restrictions beyond 12/23 (B12/23) that make the Vbeta23-RS incompatible with the Jbeta12-RS. To determine whether direct Vbeta to Jbeta joining occurs if flanking RSs are B12/23 compatible, we generated mice whose lymphocytes contained replacement of the Vbeta1412-RS with the 3'Dbeta112-RS on a TCRbeta allele lacking Dbeta segments (the Jbeta1(M6) allele). Mice heterozygous for the Jbeta1(M6) allele had dramatically increased Vbeta14(+) thymocyte and T cell numbers and decreased numbers of cells expressing other Vbetas. This altered Vbeta repertoire resulted from direct Vbeta14 to Jbeta1 rearrangements on the Jbeta1(M6) allele. Mice harboring lymphocytes homozygous for Jbeta1(M6) allele developed normal thymocyte and T cell numbers with all expressing Vbeta14. Our findings show that selective RS modifications enforce rearrangement of a specific Vbeta gene segment and demonstrate the importance of B12/23 mechanisms for ensuring generation of diverse TCRbeta repertoires.