Application of a topical anesthetic reduces pain-associated parameters during spermatic cord transection in piglet castration using a minimal anesthesia model.

OBJECTIVE: This study assessed the use of a topical anesthetic as a feasible approach to reduce pain during piglet castration using a minimal anesthesia protocol. ANIMALS: 18 male piglets, aged 3-6 days, were included in this study. METHODS: A minimal anesthetic state was induced with isoflurane administered by facemask, with anesthetic depth individually adjusted based on responses to interdigital pinch. To desensitize the scrotal skin, a vapocoolant was applied 3 times. Scrotal incisions were made subsequently and Tri-Solfen (TS) or Placebo (P) was administered in both incisional gaps. After 30 seconds, the spermatic cords were severed followed by a further application of TS/P to both incision edges. Nociception-related variables, such as mean arterial blood pressure (MAP), heart rate (HR), and nocifensive movements, were assessed. RESULTS: Significant differences in MAP changes were assessed between the TS (14 ± 4 mmHg) and the P group (36 ± 8 mmHg) for cutting the spermatic cords. Furthermore, significantly fewer nocifensive movement score points appeared in the TS than in the P group (0; IQR = 0 vs 5; IQR = 6). CLINICAL RELEVANCE: In this anesthesia model, the application of TS after skin incision significantly reduced MAP responses and nocifensive movements with spermatic cord transection compared with the application of P. However, the waiting period between TS-application and spermatic cord transection might limit the benefit of the method in conscious piglets, as pain during castration is reduced but additional stress is caused by the prolonged handling. Furthermore, using a vapocoolant did not provide anesthesia for skin incisions.

C-terminus of heat shock protein 70-interacting protein-dependent GTP cyclohydrolase I degradation in lambs with increased pulmonary blood flow.

We showed that nitric oxide (NO) signaling is decreased in the pulmonary vasculature before the development of endothelial dysfunction in a lamb model of congenital heart disease and increased pulmonary blood flow (Shunt). The elucidation of the molecular mechanism by which this occurs was the purpose of this study. Here, we demonstrate that concentrations of the endogenous NO synthase (NOS) inhibitor, asymmetric dimethylarginine (ADMA), are elevated, whereas the NOS cofactor tetrahydrobiopterin (BH(4)) is decreased in Shunt lambs. Our previous studies demonstrated that ADMA decreases heat shock protein-90 (Hsp90) chaperone activity, whereas other studies suggest that guanosine-5'-triphosphate cyclohydrolase 1 (GCH1), the rate-limiting enzyme in the generation of BH(4), may be a client protein for Hsp90. Thus, we determined whether increases in ADMA could alter GCH1 protein and activity. Our data demonstrate that ADMA decreased GCH1 protein, but not mRNA concentrations, in pulmonary arterial endothelial cells (PAECs) because of the ubiquitination and proteasome-dependent degradation of GCH1. We also found that Hsp90-GCH1 interactions were reduced, whereas the association of GCH1 with Hsp70 and the C-terminus of Hsp70-interacting protein (CHIP) increased in ADMA-exposed PAECs. The overexpression of CHIP potentiated, whereas a CHIP U-box domain mutant attenuated, ADMA-induced GCH1 degradation and reductions in cellular BH(4) concentrations. We also found in vivo that Hsp90/GCH1 interactions are decreased, whereas GCH1-Hsp70 and GCH1-CHIP interactions and GCH1 ubiquitination are increased. Finally, we found that supplementation with l-arginine restored Hsp90-GCH1 interactions and increased both BH(4) and NO(x) concentrations in Shunt lambs. In conclusion, increased concentrations of ADMA can indirectly alter NO signaling through decreased cellular BH(4) concentrations, secondary to the disruption of Hsp90-GCH1 interactions and the CHIP-dependent proteasomal degradation of GCH1.