Variability, repeatability and test-retest reliability of equine flash visual evoked potentials (FVEPs).

BACKGROUND: Visual evoked potentials (VEPs) are electrical potentials generated by neurons in the central nervous system in response to visual stimuli. A series of positive and negative wavelets in response to flash-stimuli (flash-VEP; FVEP) or reversing, iso-luminant patterns (pattern-VEP; PVEP) are recorded. Pathological conditions affecting the post-retinal pathways can alter overall waveform morphology, and also affect wavelet peak times and amplitudes. FVEPs have recently been described in horses, but more data on the variability within and between subjects is required, to adequately interpret results from clinical equine patients. Therefore, the purpose of this study was to describe the variability, repeatability and test-retest reliability of equine FVEPs in normal, adult horses. RESULTS: Equine FVEPs were recorded from one randomly selected eye in 17 horses, from both eyes in eight of these horses, and also at two separate recording sessions in six horses. N1, P2, N2 and P4 wavelets were present in 100% of the recordings in all horses, while P1, N2a, P3 and P5 were only present in some recordings. Coefficients of variation (CVs) were low for P2, N2 and P4 peak times, but higher for all amplitudes. There were no statistically significant differences comparing peak times and amplitudes between eyes or between sessions. Coefficients of repeatability (CRs) are reported for P2, N2 and P4 peak times between eyes (P2; 5 ms, N2; 18 ms, P4; 18 ms) and also between sessions (P2; 5 ms, N2; 16 ms, P4; 39 ms). Intraclass correlation coefficients (ICCs), as an estimate of test-retest reliability, was assessed to be fair to excellent for most parameters. CONCLUSIONS: This study provides important data on variability, repeatability and test-retest reliability of FVEPs in normal, adult horses. We conclude that P2, N2 and P4 peak times should be included in the evaluation of equine FVEPs. The large inherent variability of FVEP amplitudes is likely to make them less suitable and useful for establishing a diagnosis on their own in most clinical patients, but they may occasionally provide support to a clinical diagnosis.

Repeatability of the hyperglycaemic clamp for assessment of ß-cell response and insulin sensitivity in horses.

BACKGROUND: The hyperglycaemic clamp has been used for measurement of insulin sensitivity (IS) but not for measurement of ß-cell response in the horse. However, the repeatability of this test has not been reported for horses. OBJECTIVES: To determine the repeatability and reliability of measures for IS and ß-cell response to glucose using the hyperglycaemic clamp in horses. STUDY DESIGN: Repeated measures, longitudinal study. METHODS: Six healthy Standardbred mares underwent a 120-min hyperglycaemic clamp on two occasions with a 10-day washout period. Indices of repeatability and reliability were calculated from measures of IS and ß-cell response to glucose derived from each hyperglycaemic clamp. RESULTS: Measures of ß-cell response to glucose, including area under the insulin response curve in the time interval 0-120 min and during steady state (60-120 min), as well as the mean insulin concentration during steady state had coefficient of variations (CV) of 9.5, 10.5 and 9.3 respectively and intraclass correlation coefficients (ICC) of 0.93, 0.93 and 0.95 respectively. The measure of glucose disposal (M) had lower CV (12.4) and ICC (0.71) compared to measure of IS (M/I-index; CV of 17.4 and ICC of 0.89). MAIN LIMITATIONS: The study was conducted in a small number of horses. CONCLUSIONS: The hyperglycaemic clamp is a diagnostic method for assessing ß-cell response to intravenous glucose with high repeatability and reliability. Insulin sensitivity can be determined with the same test but the repeatability for these measurements is lower compared to the measurements for ß-cell response.

The effect of insulin infusion on heart rate and systemic blood pressure in horses with equine metabolic syndrome.

BACKGROUND: There is little evidence that horses with equine metabolic syndrome (EMS) have higher resting blood pressures than horses with normal insulin sensitivity. However, there are indications that EMS horses have an altered dynamic response to the cardiovascular effects of insulin. OBJECTIVES: To examine heart rate and the systemic blood pressure response in EMS and control horses during insulin infusion. STUDY DESIGN: Cross-sectional study. METHODS: Horses were examined with an oral sugar test (OST) and a euglycaemic hyperinsulinaemic clamp (EHC). Based on the results from the OST, the horses were classified as either EMS or control. Blood pressure was measured before the start of the EHC and at 60, 120 and 180 min of the EHC. RESULTS: There were 16 EMS and 12 control horses. Mean resting heart rate was higher for EMS horses compared with control (35.6 ± 5.1 vs. 30.3 ± 3.5 beats/min). Systolic, diastolic and mean arterial blood pressure did not differ between groups at the start of the EHC. Blood pressure decreased gradually during the EHC in the control group, whereas the EMS group showed no decline in systolic, diastolic or mean arterial pressure. MAIN LIMITATIONS: Limited number of horses. CONCLUSIONS: Horses with EMS have cardiovascular changes that affect resting heart rate and systemic blood pressure during insulin infusion.

Evaluation of glucose and insulin response to haylage diets with different content of nonstructural carbohydrates in 2 breeds of horses.

Information about the effect of nonstructural carbohydrates (NSCs) in forage on the postprandial glucose and insulin response in horses is scarce. This is of interest as postprandial hyperinsulinemia in horses is a risk factor for laminitis. In addition, insulin sensitivity (IS) differs between breeds. The aim was to evaluate the postprandial glucose and insulin response to haylage diets with different NSC content in horses of 2 different breeds and to evaluate the relationship between the postprandial insulin response and measures of IS derived from a frequently sampled intravenous glucose tolerance test (FSIGTT). Standardbreds (n = 9) and Icelandic horses (n = 9) with a mean body condition score of 5.5 ± 0.6 (scale 1-9) were studied. Horses were clinically healthy at the start of the study and had no history of endocrinopathic laminitis. The experiment was conducted as a replicate 3 × 3 Latin square, in which horses were fed haylage diets with low (4.2%), medium (13.6%), and high (18.2%) NSC content of dry matter. Blood sampling was performed before feeding and every 30 min until 300 min after feeding. An FSIGTT was also performed in all horses. The early (first 60 min) and the total (300 min) postprandial glucose and insulin response (area under the curve [AUC]) was higher after a meal of both medium and high NSC haylage in comparison with low NSC haylage when both breeds were combined (P ≤ 0.02). There was a main effect of breed for the early (P ≤ 0.004) but not for the total (P > 0.12) postprandial glucose and insulin response. The IS index was comparable between breeds (P = 0.75). The natural logarithm of the peak concentration, the AUC for the first 60 min and the total AUC for insulin, after a meal of medium and high NSC haylage, were moderately negatively correlated (P < 0.02; r = -0.55 to -0.72) with the natural logarithm of IS index from the FSIGTT. This relationship was not evident for haylage with low NSC content (P > 0.054). This study demonstrates that the postprandial insulin response is affected by both the NSC content of haylage and the horse's IS. However, the impact of IS was diminished when the NSC content in haylage was low (4.2% of dry matter).

Relationship Between ß-cell Response and Insulin Sensitivity in Horses based on the Oral Sugar Test and the Euglycemic Hyperinsulinemic Clamp.

BACKGROUND: A hyperbolic relationship between ß-cell response and insulin sensitivity (IS) has been described in several species including rodents, dogs, and humans. This relationship has not been elucidated in the horse. HYPOTHESIS/OBJECTIVES: To determine whether the hyperbolic relationship between ß-cell response and IS exists in horses by using indices of ß-cell response from the oral sugar test (OST) and IS measurements from the euglycemic hyperinsulinemic clamp (EHC). A second aim was to compare how well IS estimates from the OST and EHC correlate. ANIMALS: Forty-nine horses with different degrees of insulin regulation (normal-to-severe insulin dysregulation). METHODS: Cross-sectional study. Horses were examined with an OST and an EHC. RESULTS: Decreased IS was associated with increased ß-cell response in the horses. Nine of 12 comparisons between indices of ß-cell response and IS measures fulfilled the criteria for a hyperbolic relationship. Indices of IS calculated from the OST correlated highly with the insulin-dependent glucose disposal rate (M) and the insulin-dependent glucose disposal rate per unit of insulin (M/I) determined from the EHC (r = 0.81-0.87). CONCLUSIONS AND CLINICAL IMPORTANCE: A hyperbolic relationship between ß-cell response and IS exists in horses, which suggest that horses with insulin dysregulation respond not only with postprandial hyperinsulinemia but are also insulin resistant. The OST is primarily a test for ß-cell response rather than a test for IS, but calculated indices of IS from the OST may be useful to estimate IS in horses, especially when the horse is insulin resistant.

Variation in training regimens in professional showjumping yards.

REASONS FOR PERFORMING STUDY: Training regimens of showjumping horses under field conditions are largely undocumented. OBJECTIVES: The aims of this study were to quantify and compare training regimens used in professional-level showjumping yards, with respect to time exercised and type of activity. STUDY DESIGN: Prospective cohort study. METHODS: A prospective 6-month cohort study of showjumping horses in 4 European countries (The Netherlands, Sweden, Switzerland, Great Britain) was designed to analyse training and health data, in yards with several horses in training and riders competing at professional level. Riders documented the daily frequency and duration of all physical activities of the horses. Variation in training routines were compared between riders, location and time. Mixed-models analysis was used to examine factors associated with total time exercised and time spent in flatwork. RESULTS: In 4 countries, the 31 participating riders trained 263 European Warmbloods. The total days at risk (e.g. days in which the horses were considered fit for exercise) was 39,262. Mean time spent in daily exercise, including ridden work, lungeing and treadmill exercise, varied between riders from 19-52 min/day at risk. There was considerable variation in activities and level of heavy work and light exercise, i.e. turnout. Total time exercised and time spent in flatwork differed with month, country and proportion of days lost to training. Low variation of activities was associated with decreased total time trained and increased time spent in flatwork. CONCLUSIONS: Riders at this elite professional level of showjumping used training regimens that vary substantially in time spent training and other physical activities and showjumping horses are challenged differently during training despite competing at the same level. Whether all training regimens prepare the horses equally for the demands of competition remains to be determined.

Repeatability of the combined glucose-insulin tolerance test and the effect of a stressor before testing in horses of 2 breeds.

BACKGROUND: The increasing number of horses diagnosed with insulin resistance (IR) and the suggested link between IR and laminitis has highlighted the need to accurately diagnose IR in clinical practice. HYPOTHESIS/OBJECTIVES: The aim of the study was to evaluate the repeatability of the combined glucose-insulin tolerance test (CGIT) as well as to determine the effect of 2 different breeds and the effect of a stressor on the test results. ANIMALS: Clinically normal horses, 9 Standardbred horses and 9 Icelandic horses. METHODS: Prospective clinical nonrandomized trial. The CGIT was performed on all horses on 2 occasions 3 weeks apart. An additional CGIT was performed on four of the Standardbred and four of the Icelandic horses after transportation to a new environment (stressor) the day before testing. RESULTS: Calculated parameters for the glucose curve of the CGIT had low repeatability, whereas the parameters for the insulin curve had high repeatability. There was an effect of breed (Standarbred versus Icelandic horse) as well as stress on the glucose dynamics, but not on the insulin dynamics of the CGIT. CONCLUSION AND CLINICAL IMPORTANCE: Repeatability of the glucose dynamics of the CGIT is low. In addition, there appears to be breed differences in the glucose dynamics. It is therefore suggested that diagnosis of IR should not be made solely based on results from the glucose curve of the CGIT.

Expression of tlr4, md2 and cd14 in equine blood leukocytes during endotoxin infusion and in intestinal tissues from healthy horses.

The expression of tlr4, md2 and cd14 was studied in equine blood leukocytes and in intestinal samples using real time PCR. The stability of three commonly used reference genes, glyceraldehyde-3P-dehydrogenase (GAPDH), hypoxantine ribosyltransferase (HPRT) and succinate dehydrogenase complex subunit A (SDHA), was evaluated using qbase(PLUS). The equine peripheral blood mononuclear cells (eqPBMC) examined were either stimulated in vitro with Phorbol 12-myristate 13-acetate (PMA) and ionomycin or with the CpG oligodeoxynuclotide 2216 (CpG-ODN 2216) or obtained from horses before, during and after infusion of endotoxin. Intestinal tissue from healthy horses was sampled at ileum, right dorsal colon and rectum. Ranking of the three reference genes used for normalisation identified the combination HPRT/SDHA as most suitable both when determined ex vivo in leukocytes obtained from experimentally induced endotoxaemia and in eqPBMC activated in vitro while HPRT/GAPDH were most appropriate for the intestinal samples. The relative amounts of mRNA for TLR4 and MD-2 increased threefold during in vitro activation of the cells with CpG-ODN 2216 but was decreased in cultures stimulated with PMA/ionomycin. A transient elevation in the transcription of tlr4 and md2 was also evident for equine blood leukocytes following endotoxaemia. The levels of mRNA for CD14 on the other hand remained unaffected both during the induction of endotoxaemia and in the in vitro stimulated PBMCs. A low steady expression of TLR4, MD-2 and CD14 mRNA was demonstrated for the intestinal samples with no variation between the intestinal segments analysed. Thus, the foundation for real time PCR based levels of analysis of mRNA for all three components in the equine LPS receptor complex in different intestinal segments was set, making it possible to carry out future expression studies on clinical material.

Acquired equine polyneuropathy in Norway and Sweden: a clinical and epidemiological study.

REASONS FOR PERFORMING STUDY: Acquired equine polyneuropathy (AEP, also known as 'Scandinavian knuckling syndrome'), is a serious disease of unknown aetiology, which emerged clustering in horse farms in Sweden, Norway and Finland in the 1990s. Clinical and epidemiological data regarding the syndrome are scarce. OBJECTIVES: To describe the clinical and epidemiological findings and outcome in outbreaks of AEP and compare risk factors in affected and unaffected horses on affected farms in Norway and Sweden during 2007-2009. METHODS: Neurological examinations were performed and data collected regarding demography, usage, turning-out, feeding, prophylactic strategies and long-term outcome. RESULTS: Thirteen affected farms with 157 horses of various breeds, of which 42 were AEP cases, were studied. Typical digital extensor dysfunction and knuckling of pelvic limbs were noted in 34 definitive cases. Eight additional plausible cases had a severe, acute course of neurological disease. There were no signs of brain orcranial nerve dysfunction. Cases occurred from December to April, with new cases emerging within 100 days of the index case. Affected and unaffected horses were fed wrapped forage. Prevalence for AEP was 27% and case fatality 29%. The median duration of AEP in survivors was 4.4 months (1-17 months). Survivors returned to full work within 19 months (median 6.6 months). Acquired equine polyneuropathy was less prevalent in horses aged > 12 years and young horses had a higher chance of survival than older horses. Management factors did not differ between affected and unaffected horses. CONCLUSIONS: Acquired equine polyneuropathy is a potentially fatal neurological disease characterised by pelvic limb knuckling. Surviving horses returned to normal function after a long period of rest. Cases were clustered in farms during the winter/spring season. Wrapped forage was used in all farms. POTENTIAL RELEVANCE: The results provide valuable insights into the dinical examination, handling and prognosis of cases of AEP, an emerging neurological disease of unknown aetiology in horses.

Tachypnea and antipyresis in febrile horses after sedation with alpha-agonists.

BACKGROUND: Signs of tachypnea after sedation of febrile horses with alpha2-agonists have been noted previously but have not been further investigated. OBJECTIVES: To examine the effects of xylazine and detomidine on respiratory rate and rectal temperature in febrile horses and to investigate if either drug would be less likely than the other to cause changes in these variables. ANIMALS: Nine febrile horses and 9 healthy horses were included in the study. METHODS: Horses were randomly assigned to sedation with xylazine 0.5 mg/kg or detomidine 0.01 mg/kg. Heart rate and respiratory rate were recorded before sedation and at 1, 3, and 5 minutes after injection. Hourly measurements of rectal temperature were performed starting before sedation. RESULTS: All febrile horses experienced an episode of tachypnea and antipyresis after sedation. Rectal temperature in the febrile group was significantly lower at 1, 2, and 3 hours after sedation. In several measurements, the decrease was >1 degrees C. Respiratory rate in the febrile group was significantly increased after sedation. All febrile horses were breathing>40 breaths/min and 3 horses>100 breaths/min 5 minutes after sedation. No differences were noted between the 2 treatments. No significant changes in respiratory rate or temperature were noted in the reference group. CONCLUSIONS AND CLINICAL IMPORTANCE: Febrile horses can become tachypneic after sedation with detomidine or xylazine. The antipyretic properties of alpha2-agonists need consideration when evaluating patients that have been sedated several hours before examination.

Enilconazole treatment of horses with superficial Aspergillus spp. rhinitis.

BACKGROUND: Mycotic rhinitis is an uncommon disease in horses with a wide range of clinical signs and treatment options based on case studies of a single or only a few individuals. Moreover, while dogs with mycotic rhinitis appear to respond well to treatment with topical enilconazole, this approach has yet to be reported in horses. OBJECTIVES: The aim of this study was to describe the clinical findings, results of diagnostic investigations, treatment, and outcome in horses with mycotic rhinitis treated with topical enilconazole. ANIMALS: Eight horses were included in the study. METHODS: Medical records from 1995 to 2007 from an equine veterinary teaching hospital were reviewed to identify cases with a diagnosis of mycotic rhinitis. RESULTS: Eleven cases were identified, of which 8 were treated with topical enilconazole. Seven of those 8 horses recovered and had normal appearance of the nasal passages on endoscopic examination. Fungal culture, where positive, yielded only Aspergillus spp. Treatment was readily performed and no adverse effects were noted in these horses. In 5 horses, long-term follow-up revealed complete recovery for an average of 5 years. CONCLUSIONS AND CLINICAL IMPORTANCE: Mycotic rhinitis owing to Aspergillus spp. in horses can be effectively and safely treated with topical enilconazole.

Synthesis of proglycogen and macroglycogen in skeletal muscle of standardbred trotters after intermittent exercise.

REASONS FOR PERFORMING STUDY: The degradation of glycogen and its two forms, proglycogen (PG) and macroglycogen (MG) has been studied in horses performing different types of exercise, but no information is available about the resynthesis of PG and MG after exercise. OBJECTIVES: To determine the resynthesis of PG and MG in skeletal muscle after intermittent uphill exercise. METHODS: At a training camp 9 well-trained Standardbred trotters performed a training session comprising a warm-up period, 7 repeated 500 m bouts of exercise on an uphill slope and a recovery period. Muscle biopsies (m. gluteus medius) for analysis of PG, MG, glucose and glucose-6-phosphate were taken at rest, at the end of exercise and 1, 4, 8, 24, 48 and 72 h post exercise. Blood samples for analysis of glucose, lactate and insulin were collected before exercise, immediately after the last bout of exercise and then as for the muscle biopsies. RESULTS: The MG and PG concentration pre-exercise was 311 - 47 and 305 +/- 55 mmol/kg dwt respectively. The exercise caused a decrease in PG (A 63 +/- 26 mmol/kg dwt) and MG (delta 136 +/- 68 mmol/kg dwt). Immediately after the last sprint plasma glucose and lactate increased compared to values pre-exercise. During the first hour post exercise there was a further decrease in MG in 7 out of 9 horses. The rate of glycogen resynthesis during 1-24 h was higher for MG than for PG. The rate of muscle glycogen resynthesis thereafter was slower and did not differ between MG and PG up to 72 h. CONCLUSION: After repeated bouts of exercise on a slope, resynthesis of glycogen is a slow process and the resynthesis of proglycogen differs from that of macroglycogen. The fraction most depleted during exercise (MG) had no resynthesis during the first hour of recovery but then had the highest rate of resynthesis during the remainder of the first 24 h period. POTENTIAL RELEVANCE: If the time between exercise sessions during training is too short the recovery period will be inadequate for complete restoration of muscle glycogen.

Pro- and macroglycogenolysis in skeletal muscle during maximal treadmill exercise.

The purpose was to investigate the degradation of proglycogen and macroglycogen in skeletal muscle during intense exercise. Ten Standardbred trotters performed a maximal treadmill exercise test comprising a warm-up period, an exercise period, starting at 7 m/s with increments of 1 m/s every 60 s until the onset of fatigue (mean +/- s.d. 246 +/- 32 s) and a walking recovery period. Muscle biopsies were taken at rest, immediately after exercise and 15 min postexercise. The exercise caused a marked anaerobic metabolism as shown by the decrease in both muscle ATP and creatine phosphate and increase in muscle lactate. Free muscle glucose increased immediately postexercise and a further increase was noted 15 min later. There was a significant decrease (P<0.05) in proglycogen (57.1 +/- 22.2 mmol/kg dw) and macroglycogen (63.0 +/- 65.5 mmol/kg dw) during exercise. The proglycogen concentration tended to increase 15 min after exercise (19.9 +/- 27.3 mmol/kg dw; P = 0.06). The results from this study demonstrate that both proglycogen and macroglycogen contribute equally to glycogenolysis during intense exercise and suggest that glycogen resynthesis starts in the proglycogen pool.

Halicephalobus gingivalis encephalomyelitis in a horse.

An 8-year-old, Arabian mare presented with acute progressive ataxia and a firm swelling over the right mandible. Radiographs revealed multiple radiolucent areas on the mandibles. The mare's neurological signs progressed, she was consequently euthanized. Postmortem examination revealed mandibular granulomatous reactions and meningoencephalitis due to the nematode Halicephalobus gingivalis.