Measurement of C‐peptide concentrations and responses to somatostatin,glucose infusion,and insulin resistance in horses

Reasons for performing study: Hyperinsulinaemia is detected in horses with insulin resistance (IR) and has previously been attributed to increased pancreatic insulin secretion. Connecting peptide (C‐peptide) can be measured to assess pancreatic function because it is secreted in equimolar amounts with insulin and does not undergo hepatic clearance. Hypothesis: A human double antibody radioimmunoassay (RIA) detects C‐peptide in equine serum and concentrations would reflect responses to different stimuli and conditions. Methods: A validation procedure was performed to assess the RIA. Six mature mares were selected and somatostatin administered i.v. as a primed continuous rate infusion, followed by 50 nmol human C‐peptide i.v. Insulin and C‐peptide concentrations were measured in horses (n = 6) undergoing an insulin‐modified frequently sampled i.v. glucose tolerance test, and in horses with insulin resistance (n = 10) or normal insulin sensitivity (n = 20). Results: A human RIA was validated for use with equine sera. Endogenous C‐peptide secretion was suppressed by somatostatin and median (range) clearance rate was 0.83 (0.15–1.61) ml/min/kg bwt. Mean ± s.d. C‐peptide‐to‐insulin ratio significantly (P = 0.004) decreased during the glucose tolerance test from 3.60 ± 1.95 prior to infusion to 1.03 ± 0.18 during the first 20 min following dextrose administration. Median C‐peptide and insulin concentrations were 1.5‐ and 9.5‐fold higher, respectively in horses with IR, compared with healthy horses. Conclusions: Endogenous C‐peptide secretion decreases in response to somatostatin and increases after dextrose infusion. Results suggest that relative insulin clearance decreases as pancreatic secretion increases in response to dextrose infusion. Hyperinsulinaemia in insulin resistant horses may be associated with both increased insulin secretion and decreased insulin clearance. Potential relevance: Both C‐peptide and insulin concentrations should be measured to assess pancreatic secretion and insulin clearance in horses.     

Effect of Different Combinations of Cryoprotectants on In Vitro Maturation of Immature Buffalo (Bubalus bubalis) Oocytes Vitrified by Straw and Open‐Pulled Straw Methods

This study was designed to evaluate effects of different combinations of cryoprotectants on the ability of vitrified immature buffalo oocytes to undergo in vitro maturation. Straw and open‐pulled straw (OPS) methods for vitrification of oocytes at the germinal vesicle stage also were compared. The immature oocytes were harvested from ovaries of slaughtered animals and were divided into three groups: (i) untreated (control); (ii) exposed to cryoprotectant agents (CPAs); or (iii) cryopreserved by straw and OPS vitrification methods. The vitrification solution (VS) consisted of 6 m ethylene glycol (EG) as the standard, control vitrification treatment, and this was compared with 3 m EG + 3 m dimethyl sulfoxide (DMSO), 3 m EG + 3 m glycerol, and 3 m DMSO + 3 m glycerol. Cryoprotectants were added in two steps, with the first step concentration half that of the second (and final) step concentration. After warming, oocyte samples were matured by standard methods and then fixed and stained for nuclear evaluation. Rates of MII oocytes exposed to CPAs without vitrification were lower (54.3 ± 1.9% in EG, 47.5 ± 3.4% in EG + DMSO, 36.8 ± 1.2% in EG + glycerol and 29.9 ± 1.0% in DMSO + glycerol; p < 0.05) than for the control group (79.8 ± 1.3%). For all treatments in each vitrification experiment, results were nearly identical for straws and OPS, so all results presented are the average of these two containers. The percentages of oocytes reaching telophase‐I or metaphase‐II stages were lower in oocytes cryopreserved using all treatments when compared with control. However, among the vitrified oocytes, the highest maturation rate was seen in oocytes vitrified in EG + DMSO (41.5 ± 0.6%). Oocytes cryopreserved in all groups with glycerol had an overall low maturation rate 19.0 ± 0.6% for EG + glycerol and 17.0 ± 1.1% for DMSO + glycerol. We conclude that the function of oocytes was severely affected by both vitrification and exposure to cryoprotectants without vitrification; the best combination of cryoprotectants was EG + DMSO for vitrification of immature buffalo oocytes using either straw or OPS methods.