Effect of exercise, training, circadian rhythm, age, and sex on insulin-like growth factor-1 in the horse.

Insulin-like growth factor-1 could be a useful marker in the horse for diagnostic, selection, or forensic purposes, provided its physiological regulation is well understood. The objective of this study was to investigate factors, such as acute exercise, fitness training, time of day, sex, and age, that may influence serum IGF-1 in normal, healthy horses. Throughout a 9-wk training program, 6 geldings maintained a mean (+/- SEM) IGF-1 concentration of 302 +/- 29 ng/mL. Moderate or high intensity exercise had no effect on IGF-1 concentrations, when pre- and postexercise values were compared. Over a 24-h period, there was some variation in IGF-1 concentrations but no clear diurnal rhythm. Concentrations of IGF-1 were measured in a large population of thoroughbred horses (1,880) on 3 continents. The population deviated slightly from a normal distribution (P < 0.001) because of large IGF-1 concentrations in 10 horses. The global mean IGF-1 concentration was 310 +/- 2.2 ng/mL, with a greater mean value (P < 0.001) in gonad-intact males (336 +/- 5.6 ng/mL) than in females (303 +/- 3.2 ng/mL) or geldings (302 +/- 3.2 ng/mL). However, the greatest IGF-1 concentrations observed for all stallions, mares, and geldings were 627, 676, and 709 ng/mL, respectively. In mares and geldings, IGF-1 concentrations showed a gradual decrease with advancing age (P < 0.001), but the effect was much less marked in stallions. This study confirms that IGF-1 concentrations are stable, compared with GH concentrations, in the horse and that a meaningful measure of IGF-1 status can be obtained from a daily serum sample.

Confirmed Datura poisoning in a horse most probably due to D. ferox in contaminated tef hay.

Two out of a group of 23 mares exposed to tef hay contaminated with Datura ferox (and possibly D. stramonium) developed colic. The 1st animal was unresponsive to conservative treatment, underwent surgery for severe intestinal atony and had to be euthanased. The 2nd was less seriously affected, responded well to analgesics and made an uneventful recovery. This horse exhibited marked mydriasis on the first 2 days of being poisoned and showed protracted, milder mydriasis for a further 7 days. Scopolamine was chemically confirmed in urine from this horse for 3 days following the colic attack, while atropine could just be detected for 2 days. Scopolamine was also the main tropane alkaloid found in the contaminating plant material, confirming that this had most probably been a case of D. ferox poisoning. Although Datura intoxication of horses from contaminated hay was suspected previously, this is the 1st case where the intoxication could be confirmed by urine analysis for tropane alkaloids. Extraction and detection methods for atropine and scopolamine in urine are described employing enzymatic hydrolysis followed by liquid-liquid extraction and liquid chromatography tandem mass spectrometry (LC/MS/MS).

Administration of bovine, porcine and equine growth hormone to the horse: effect on insulin-like growth factor-I and selected IGF binding proteins.

This study investigated the biochemical effects of administration of three types of recombinant growth hormone (GH; somatotropin) to the Thoroughbred horse. Equine or bovine or porcine GH was administered at a recommended dosage to 3-5-year old Thoroughbred geldings, for up to 21 days. It was shown that, in addition to equine GH, bovine and porcine GH were active in the horse; however, porcine GH caused injection-site reactions that were so serious that administration had to be terminated. The concentrations of a range of GH-related serum protein markers were determined before, during and after the administration period. Because of the short half-life of GH itself, the objective was to identify GH-related markers that showed changes in concentration and which could be used as indicators of the abuse of these hormones. Among the possible markers identified, serum total insulin-like growth factor (IGF)-I was shown to be the most promising, increasing to 270% of the basal concentration for equine GH administration. After GH administration, IGF-I took longer to attain baseline concentrations than the time required for GH concentrations to recover to normal. The concentration obtained from the administration significantly exceeded natural concentrations for IGF-I, as was determined from a population of more than 2000 Thoroughbred horses in three continents. The concentrations of serum free IGF-I and IGF binding protein 3 (IGFBP-3) were also shown to be significantly affected by equine and bovine GH.

Growth hormone abuse in the horse: preliminary assessment of a mass spectrometric procedure for IGF-1 identification and quantitation.

Previous studies have shown that insulin-like growth factor 1 (IGF-1) is a promising marker for the detection of growth hormone (GH) abuse in the horse. The significant increases observed with GH administration in comparison to natural levels imply the possibility of setting a threshold level for IGF-1 that would be indicative of GH abuse. Although an immunoradiometric assay (IRMA) has been identified as a reliable screening method, a more specific IGF-1 quantification method needs to be developed for the prosecution of GH abuse by horseracing authorities. This study describes such an HPLC electrospray mass spectrometry (LC/ESI-MS) method that was developed and then assessed for the specific analysis of IGF-1 at the low levels encountered in serum. The structural identity of IGF-1 was confirmed by endoproteinase Asp-N digestion followed by LC/MS and LC/MS/MS characterisation. This was followed by quantification of IGF-1 as the intact molecule against an internal standard.

Mechanism for the stabilization in vivo of the aziridine precursor --(4-acetoxyphenyl)-2-chloro-N-methyl-ethylammonium chloride by serum proteins.

Oral and intraperitoneal administration of 2-(4-acetoxyphenyl)-2-chloro-N-methyl-ethylammonium chloride (Compound A), an analogue of phenyl aziridine precursors that occur in the shrub Salsola tuberculatiformis Botsch, had a contraceptive effect on female Wistar rats with a concomitant decrease in total body, uterus, and every mass and an increase in abronal mass. Compound A elicited a Type II difference spectrum and inhibited the Type I deoxycorticosterone (DOC) induced difference spectrum of sheep adrenal cytochrome P450c11 in a manner similar to that of S2, a biologically active fraction isolated from S. tuberculatiformis. The effects of Compound A on the spectral properties of P450c11 were diminished with time in PBS. Electrospray mass spectrometry (ES-MS) indicated that the rate of cyclization of Compound A to the corresponding aziridine followed a time course similar to the attenuation of cytochrome P450c11 inhibition. It was concluded that the aziridine precursor. Compound A, rather than aziridine itself, was the inhibiting agent of sheep adrenal P450c11. Addition of sheep and rat plasma prevented the attenuation of the effect of Compound A on the spectral properties of cytochrome P450c11. Subsequent ES-MS analysis indicated that Compound A was stabilized in plasma by sex hormone binding globulin and corticosteroid binding globulin. These results suggest a mechanism whereby natural plant products, which are highly reactive and unstable in vitro, can be stabilized by binding to plasma proteins, and so remain biologically active in vivo.