Intervertebral disc degeneration in warmblood horses: Histological and biochemical characterization.

Gross morphology of healthy and degenerated intervertebral discs (IVDs) is largely similar in horses as in dogs and humans. For further comparison, the biochemical composition and the histological and biochemical changes with age and degeneration were analyzed in 41 warmblood horses. From 33 horses, 139 discs and 2 fetal vertebral columns were evaluated and scored histologically. From 13 horses, 73 IVDs were assessed for hydration, DNA, glycosaminoglycans, total collagen, hydroxyl-lysyl-pyridinoline, hydroxylysine, and advanced glycation end-product (AGE) content. From 7 horses, 20 discs were assessed for aggrecan, fibronectin, and collagen type 1 and 2 content. Histologically, tearing of the nucleus pulposus (NP) and cervical annulus fibrosus (AF), and total histological score (tearing and vascular proliferation of the AF, and chondroid metaplasia, chondrocyte-like cell proliferation, presence of notochordal cells, matrix staining, and tearing of the NP) correlated with gross degeneration. Notochordal cells were not seen in IVDs of horses. Age and gross degeneration were positively correlated with AGEs and a fibrotic phenotype, explaining gross degenerative changes. In contrast to dogs and humans, there was no consistent difference in glycosaminoglycan content and hydration between AF and NP, nor decrease of these variables with age or degeneration. Hydroxylysine decrease and collagen 1 and AGEs increase were most prominent in the NP, suggesting degeneration started in the AP. In caudal cervical NPs, AGE deposition was significantly increased in grossly normal IVDs and total collagen significantly increased with age, suggesting increased biomechanical stress and likelihood for spinal disease in this part of the vertebral column.

Equine cervical intervertebral disc degeneration is associated with location and MRI features.

Morphology of the equine cervical intervertebral disc is different from that in humans and small companion animals and published imaging data are scarcely available. The objectives of this exploratory, methods comparison study were (a) to describe MRI features of macroscopically nondegenerated and degenerated intervertebral discs (b) to test associations between spinal location and macroscopic degeneration or MRI-detected annular protrusion and between MRI-detected annular protrusion and macroscopic degeneration, and (c) to define MRI sequences for characterizing equine cervical intervertebral disc degeneration. Ex vivo MRI of intervertebral discs was performed in 11 horses with clinical signs related to the cervical region prior to macroscopic assessment. Mixed-effect logistic regression modeling included spinal location, MRI-detected annular protrusion, and presence of macroscopic degeneration with "horse" as random effect. Odds ratio and 95% confidence interval were determined. Reduced signal intensity in proton density turbo SE represented intervertebral disc degeneration. Signal voids due to presence of gas and/or hemorrhage were seen in gradient echo sequences. Presence of macroscopic intervertebral disc degeneration was significantly associated with spinal location with odds being higher in the caudal (C5 to T1) versus cranial (C2 to C5) part of the cervical vertebral column. Intervertebral discs with MRI-detected annular protrusion grades 2-4 did have higher odds than with grade 1 to have macroscopic degeneration. It was concluded that MRI findings corresponded well with gross macroscopic data. Magnetic resonance imaging of the equine cervical intervertebral disc seems to be a promising technique, but its potential clinical value for live horses needs to be explored further in a larger and more diverse population of horses.

Intervertebral Disc Degeneration in Warmblood Horses: Morphology, Grading, and Distribution of Lesions.

Equine intervertebral disc degeneration is thought to be rare and of limited clinical relevance, although research is lacking. To objectively assess pathological changes of the equine intervertebral disc and their clinical relevance, description of the normal morphology and a practical, biologically credible grading scheme are needed. The objectives of this study are to describe the gross and histological appearance of the equine intervertebral discs and to propose a grading scheme for macroscopic degeneration. Spinal units from 33 warmblood horses were grossly analyzed and scored. Of the 286 intervertebral discs analyzed, 107 (37%) were assigned grade 1 and grade 2 (considered normal) and were analyzed histologically. A nucleus pulposus and an annulus fibrosus could be identified macroscopically and histologically. Histologically, the nucleus pulposus was composed of a cartilaginous matrix and the annulus fibrosus of parallel collagenous bands. A transition zone was also histologically visible. Intra- and inter-observer reliability scores were high for all observers. Higher grades were associated with greater age. Gross changes associated with equine intervertebral disc degeneration (grades 3-5)-that is, yellow discoloration, cleft formation (tearing), and changes in consistency of the nucleus pulposus-were largely similar to those in humans and dogs and were most prevalent in the caudal cervical spine. Equine intervertebral disc degeneration was not associated with osteophyte formation. Changes of the vertebral bone were most common in the thoracolumbar spine but were not correlated with higher grades of intervertebral disc degeneration. Thus, changes of the vertebral bone should be excluded from grading for equine intervertebral disc degeneration.

Copper-associated hepatic cirrhosis in a Friesian horse.

A 6-year-old Friesian stallion was examined because of signs of exercise intolerance, stiff gait and symmetrical hind weakness, and increased serum liver enzymes. On presentation, the horse showed muscle atrophy of the hindquarters. Neurological investigation showed no abnormalities. Laboratory findings revealed a prolonged prothrombin time and increased levels of alkaline phosphatase (AF), aspartate aminotransferase (ASAT), gamma-glutamyl-transferase (GGT), lactate dehydrogenase (LDH), and bile acids. Histological evaluation of the liver revealed severe cirrhosis and intracytoplasmic greyish brown granules in almost all hepatocytes, sinusoidal Kuppfer cells, and macrophages. These granules stained strongly for copper. Treatment to slow hepatic fibrosis was advised and included oral prednisolone administration for at least 1 month. A diet to support liver function was formulated by a nutritional specialist, and vitamin E was advised as dietary supplement to support neuromuscular function. Soon after diagnosis, the animal showed signs of intravascular haemolysis, with the presence of Heinz bodies in peripheral blood smears, and haemoglobinuria. On the basis of this haemolytic crisis and the poor prognosis of the chronic hepatic disease, the horse was euthanized at the owners' request. Although we could not establish the cause of the hepatic copper accumulation, this case report highlights that excessive copper in the liver should be considered in the differential diagnosis of hepatic cirrhosis and Heinz body anaemia in the horse.

Direct detection of SARS-CoV-2 antisense and sense genomic RNA in human saliva by semi-autonomous fluorescence in situ hybridization: A proxy for contagiousness?

Saliva is a matrix which may act as a vector for pathogen transmission and may serve as a possible proxy for SARS-CoV-2 contagiousness. Therefore, the possibility of detection of intracellular SARS-CoV-2 in saliva by means of fluorescence in situ hybridization is tested, utilizing probes targeting the antisense or sense genomic RNA of SARS-CoV-2. This method was applied in a pilot study with saliva samples collected from healthy persons and those presenting with mild or moderate COVID-19 symptoms. In all participants, saliva appeared a suitable matrix for the detection of SARS-CoV-2. Among the healthy, mild COVID-19-symptomatic and moderate COVID-19-symptomatic persons, 0%, 90% and 100% tested positive for SARS-CoV-2, respectively. Moreover, the procedure allows for simultaneous measurement of viral load ('presence', sense genomic SARS-CoV-2 RNA) and viral replication ('activity', antisense genomic SARS-CoV-2 RNA) and may yield qualitative results. In addition, the visualization of DNA in the cells in saliva provides an additional cytological context to the validity and interpretability of the test results. The method described in this pilot study may be a valuable diagnostic tool for detection of SARS-CoV-2, distinguishing between 'presence' (viral load) and 'activity' (viral replication) of the virus. Moreover, the method potentially gives more information about possible contagiousness.

Effect of a long-term high-energy diet on cardiovascular parameters in Shetland pony mares.

BACKGROUND: Changes in cardiovascular parameters, including blood pressure (BP) and cardiac anatomical dimensions, are an inconsistent feature of the equine metabolic syndrome. The order in which these changes arise is unknown. OBJECTIVES: Determine the order in which EMS-associated changes in cardiovascular parameters arise. ANIMALS: Twenty Shetland pony mares. METHODS: High-energy (HE) diet mares were fed 200% of net energy requirements for 1 (n = 3) or 2 (n = 7) consecutive diet-years, with 17 weeks of hay-only between years. Noninvasive BP measurements and echocardiograms were performed during both years. Resting 24-hour ECGs and measurements of autonomic tone (splenic volume and packed cell volume [PCV]) were performed at the end of diet-year 1. Results were compared to control mares receiving a maintenance diet for 1 (n = 7) or 2 (n = 3) consecutive years. RESULTS: In year 1, HE mares had significantly higher values than control mares for mean relative left ventricular wall thickness (P = .001). After 2 diet-years, mean systolic (P = .003), diastolic (P < .001) and mean arterial BP (P = .001), heart rate (HR; P < .001), and mean left ventricular wall thickness (P = .001) also were significantly increased in HE compared to control mares. No pathological arrhythmias or differences in splenic volume or PCV were detected. CONCLUSIONS AND CLINICAL IMPORTANCE: Ingesting a HE diet first induced minor changes in BP, and progressed to left-sided cardiac hypertrophy in Shetland pony mares. These findings are of interest given the increasing incidence of obesity in horses.

Cervical articular process joint osteochondrosis in Warmblood foals.

BACKGROUND: In Warmblood horses, degenerative joint disease is involved in cervical malformation and malarticulation (CVM). The degree of contribution of articular process joint (APJ) osteochondrosis (OC) is not clear. OBJECTIVES: (a) To explore the presence of predilection sites for APJ OC in cervical and cranial thoracic vertebral columns of Warmblood foals and (b) to examine the correlation of such a site with the predilection site of CVM. STUDY DESIGN: Case series. METHODS: Seven hundred APJ facets of C2 to T2 of 29 foals (11 months gestation to 12 months [median age 7 days; range 365 days; 95% confidence interval [95% CI] 2-47 days]) were examined for OC and prevalence between joints, and the predilection site for CVM and the cranial cervical vertebral column were evaluated. RESULTS: About 20.6% of facets revealed OC. There was no predilection site. Prevalence decreased with age up to 1 year (odds ratio [OR] 0.997; (95% CI 0.975-0.998)) but not up to 5 months. Severity increased with age in all age ranges (up to 1 year OR 1.023; 95% CI 1.005-1.049; >1-5 months, OR 1.203; 95% CI 1.014e+00-1.921; up to 1 month, OR 1.114; 95% CI 1.041-1.228). Highest prevalence was in cranial facets of the cervical and cervical-thoracic joints and in caudal facets of the thoracic joint up to 1 year and up to 1 month (OR 0.364; 95% CI 0.170-0.745, OR 0.434; 95% CI: 0.235-0.782, OR 7.665; 95% CI: 1.615-66.553 and OR 0.400; 95% CI 0.170-0.880, OR 0.351; 95% CI 0.172-0.700, OR 5.317; 95% CI 1.098-44.344 respectively). MAIN LIMITATIONS: Two-thirds of the foals were less than 1 month of age. CONCLUSIONS: Articular process joint OC in Warmblood foals is common and is not more prevalent at CVM predilection sites, suggesting that abnormalities of enchondral ossification may not be major contributors to CVM.

Effects of acute exercise and long-term exercise on total Na+,K+ -ATPase content and Na+,K+ -ATPase isoform expression profile in equine muscle.

OBJECTIVE: To investigate the effects of acute exercise and long-term training on Na(+),K(+)-ATPase content, mRNA isoforms, and protein concentration in equine muscle. ANIMALS: 6 Standardbreds. PROCEDURES: Horses performed a bout of exercise on a treadmill before and after 18 weeks of combined interval and endurance training. Muscle biopsy specimens were obtained from vastus lateralis muscle (VLM) and pectoralis descendens muscle (PDM) before and after exercise. The Na(+),K(+)-ATPase content, mRNA isoforms, and protein concentrations were determined by use of [(3)H]ouabain binding, real-time PCR assay, and western blotting, respectively. RESULTS: 6 Na(+),K(+)-ATPase mRNA isoforms were present in equine muscle, but only A2 and B1 proteins were detected. Exercise before training resulted in increases of mRNA isoforms A1, A2, A3, and B2 in VLM and A1 and B3 in PDM. Training increased resting values for mRNA isoforms A3 and B1 in VLM and B3 in PDM. The Na(+),K(+)-ATPase, [(3)H]ouabain binding, and proteins of mRNA A2 and B1 increased in VLM, whereas in PDM, only A2 protein increased as a result of training. After training, effects of strenuous exercise on mRNA expression were no longer detectable. CONCLUSIONS AND CLINICAL RELEVANCE: Equine muscle contained all Na(+),K(+)-ATPase mRNA isoforms, but only A2 and B1 proteins could be detected. Expression of these isoforms changed as a result of strenuous exercise and long-term training, representing an adaptive response. Determination of Na(+),K(+)-ATPase gene expression may be relevant for understanding alterations in excitability during neuromuscular diseases.

(Over)training effects on quantitative electromyography and muscle enzyme activities in standardbred horses.

Too intensive training may lead to overreaching or overtraining. To study whether quantitative needle electromyography (QEMG) is more sensitive to detect training (mal)adaptation than muscle enzyme activities, 12 standardbred geldings trained for 32 wk in age-, breed-, and sex-matched fixed pairs. After a habituation and normal training (NT) phase (phases 1 and 2, 4 and 18 wk, respectively), with increasing intensity and duration and frequency of training sessions, an intensified training (IT) group (phase 3, 6 wk) and a control group (which continued training as in the last week of phase 2) were formed. Thereafter, all horses entered a reduced training phase (phase 4, 4 wk). One hour before a standardized exercise test (SET; treadmill), QEMG analysis and biochemical enzyme activity were performed in muscle or in biopsies from vastus lateralis and pectoralis descendens muscle in order to identify causes of changes in exercise performance and eventual (mal)adaptation in skeletal muscle. NT resulted in a significant adaptation of QEMG parameters, whereas in muscle biopsies hexokinase activity was significantly decreased. Compared with NT controls, IT induced a stronger adaptation (e.g., higher amplitude, shorter duration, and fewer turns) in QEMG variables resembling potentially synchronization of individual motor unit fiber action potentials. Despite a 19% decrease in performance of the SET after IT, enzyme activities of 3-hydroxyacyl dehydrogenase and citrate synthase displayed similar increases in control and IT animals. We conclude that 1) QEMG analysis is a more sensitive tool to monitor training adaptation than muscle enzyme activities but does not discriminate between overreaching and normal training adaptations at this training level and 2) the decreased performance as noted in this study after IT originates most likely from a central (brain) rather than peripheral level.

Plasma acylcarnitine and fatty acid profiles during exercise and training in Standardbreds.

OBJECTIVE: To evaluate alterations in skeletal muscle carnitine metabolism during exercise and training by measuring changes in plasma acylcarnitine concentrations in Standardbreds. ANIMALS: 10 Standardbred geldings with a mean +/- SD age of 20 +/- 2 months and weight of 384 +/- 42 kg. PROCEDURES: In a 32-week longitudinal study, training on a treadmill was divided into 4 phases as follows: phase 1, acclimatization for 4 weeks; phase 2, 18 weeks with alternating endurance and high-intensity exercise training; phase 3, increased training volume and intensity for another 6 weeks; and phase 4, deconditioning for 4 weeks. In phase 3, horses were randomly assigned to 2 groups as follows: control horses (which continued training at the same level as in phase 2) and high-intensity exercise trained horses. At the end of each phase, a standardized exercise test (SET) was performed. Plasma acylcarnitine, fatty acids, and lactic acid and serum beta-hydroxybutyric acid (BHBA) concentrations were assessed before and at different time points after each SET. RESULTS: Plasma lactic acid, total nonesterified fatty acids, 3-hydroxyisobutyric acid, and acetylcarnitine (C2-carnitine) concentrations significantly increased during SETs, whereas serum BHBA, plasma propionylcarnitine (C3-carnitine), and plasma butyryl- and isobutyrylcarnitine (C4-carnitine) concentrations decreased significantly, compared with those before SETs. CONCLUSIONS AND CLINICAL RELEVANCE: Our findings indicated that the plasma acylcarnitine profile in horses likely reflects skeletal muscle carnitine metabolism following exercise, thereby providing a possible practical method to investigate potential disorders in carnitine metabolism in horses with myopathy.

Effect of exercise on activation of the p38 mitogen-activated protein kinase pathway, c-Jun NH2 terminal kinase, and heat shock protein 27 in equine skeletal muscle.

OBJECTIVE: To investigate the effects of exercise on activation of mitogen-activated protein kinase (MAPK) signaling proteins in horses. ANIMALS: 6 young trained Standardbred geldings. PROCEDURE: Horses performed a 20-minute bout of exercise on a treadmill at 80% of maximal heart rate. Muscle biopsy specimens were obtained from the vastus lateralis and pectoralis descendens muscles before and after exercise. Amount of expression and intracellular location of phosphospecific MAPK pathway intermediates were determined by use of western blotting and immunofluorescence staining. RESULTS: Exercise resulted in a significant increase in phosphorylation of p38 pathway intermediates, c-Jun NH2 terminal kinase (JNK), and heat shock protein 27 (HSP27) in the vastus lateralis muscle, whereas no significant changes were found in phosphorylation of extracellular regulated kinase. In the pectoralis descendens muscle, phosphorylation of p38 and HSP27 was significantly increased after exercise. Immunohistochemical analysis revealed fiber-type- specific locations of phosphorylated JNK in type 2a/b intermediate and 2b fibers and phosphorylated p38 in type 1 fibers. Phosphorylated HSP27 was strongly increased after exercise in type 1 and 2a fibers. CONCLUSIONS AND CLINICAL RELEVANCE: The p38 pathway and JNK are activated in the vastus lateralis muscle after a single 20-minute bout of submaximal exercise in trained horses. Phosphorylation of HSP27 as detected in the study reported here is most likely induced through the p38 signaling pathway.

The potential and limitations of quantitative electromyography in equine medicine.

This review discusses the scope of using (quantitative) electromyography (EMG) in diagnosing myopathies and neuropathies in equine patients. In human medicine, many EMG methods are available for the diagnosis, pathophysiological description and evaluation, monitoring, or rehabilitation of patients, and some of these techniques have also been applied to horses. EMG results are usually combined with other neurophysiological data, ultrasound, histochemistry, biochemistry of muscle biopsies, and clinical signs in order to provide a complete picture of the condition and its clinical course. EMG technology is commonly used in human medicine and has been subject to constant development and refinement since its introduction in 1929, but the usefulness of the technique in equine medicine is not yet widely acknowledged. The possibilities and limitations of some EMG applications for equine use are discussed.

Evaluation of glucose tolerance and intestinal luminal membrane glucose transporter function in horses with equine motor neuron disease.

OBJECTIVE: To confirm whether the plasma glucose concentration curve obtained during oral glucose tolerance tests (OGTTs) in horses with equine motor neuron disease (EMND) is decreased, compared with that obtained in clinically normal horses, and determine whether that decrease is a result of defective glucose metabolism or intestinal glucose transport dysfunction. ANIMALS: 8 horses with EMND and 44 matched control horses. PROCEDURE: Electromyography and OGTTs were performed in all 8 affected horses and 10 control horses. Intravenous GTTs (IVGTTs) were performed in 6 affected horses and another 11 control horses. The activity and levels of jejunal luminal membrane glucose transporter (Na+ / glucose cotransporter isoform 1 [SGLT1]) were measured in 2 affected horses and 23 control horses. RESULTS: In horses with EMND, generalized neuropathy was detected via quantitative electromyography; the mean increase in plasma glucose concentration during the OGTT was significantly decreased, compared with the value in control horses. During the IVGTT the mean increase in plasma glucose concentration was significantly lower than that of control horses. The activity and levels of SGLT1 in 2 affected horses were similar to those of control horses. Diagnosis of EMND was confirmed postmortem in all affected horses. CONCLUSIONS AND CLINICAL RELEVANCE: Data suggest that the decreased plasma glucose curve obtained in horses with EMND during OGTTs (compared with control horses) is a result of overall enhanced glucose metabolism or abnormalities in the facilitated glucose transporters; definitive identification of the underlying mechanisms could aid in the development of appropriate treatments of EMND in horses.

Evaluation of glucose metabolism in three horses with lower motor neuron degeneration.

OBJECTIVES: To determine whether increased glucose metabolism is the potential cause of the decreased plasma glucose curve determined after oral glucose tolerance testing in horses with lower motor neuron degeneration. ANIMALS: 3 horses with signs suggestive of lower motor neuron degeneration, 1 horse with malignant melanoma with multiple metastases, and an obese but otherwise healthy horse. Procedures-Glucose metabolism was assessed by use of the hyperglycemic clamp and euglycemic hyperinsulinemic clamp techniques. RESULTS: Mean rate of glucose metabolism of horses with lower motor neuron degeneration was significantly greater (mean, 3.7 times greater than control horses; range, 2.1 to 4.8 times greater) than that reported in 5 healthy control horses (41 +/- 13 micromol/kg/min vs 11 +/- 4.5 micromol/kg/min, respectively). In addition, one of the affected horses, an 8-year-old warmblood gelding, had a 5.6-times increased sensitivity to exogenously administered insulin, compared with that reported in 5 healthy control horses. Pancreatic insulin secretion was not insufficient in horses with lower motor neuron degeneration. Findings in the 2 diseased control horses were unremarkable. CONCLUSIONS AND CLINICAL RELEVANCE: Increased glucose metabolism in horses with lower motor neuron degeneration may be the cause of the decreased plasma glucose curve detected after oral glucose tolerance testing. This finding could aid in developing supportive treatments with respect to adequate glucose and vitamin E supplementation.

Effects of short- and long-term recombinant equine growth hormone and short-term hydrocortisone administration on tissue sensitivity to insulin in horses.

OBJECTIVE: To determine the effects of short-term IV administration of hydrocortisone or equine growth hormone (eGH) or long-term IM administration of eGH to horses on tissue sensitivity to exogenous insulin. ANIMALS: 5 Standardbreds and 4 Dutch Warmblood horses. PROCEDURE: The euglycemic-hyperinsulinemic clamp technique was used to examine sensitivity of peripheral tissues to exogenous insulin 24 hours after administration of a single dose of hydrocortisone (0.06 mg/kg), eGH (20 microg/kg), or saline (0.9% NaCl) solution and after long-term administration (11 to 15 days) of eGH to horses. The amounts of metabolized glucose (M) and plasma insulin concentration (I) were determined. RESULTS: Values for M and the M-to-I ratio were significantly higher 24 hours after administration of a single dose of hydrocortisone than after single-dose administration of eGH or saline solution. After long-term administration of eGH, basal I concentration was increased and the mean M-to-I ratio was 22% lower, compared with values for horses treated with saline solution. CONCLUSIONS AND CLINICAL RELEVANCE: Increases in M and the M-to-I ratio after a single dose of hydrocortisone imply that short-term hydrocortisone treatment increases glucose use by, and insulin sensitivity of, peripheral tissues. Assuming a single dose of hydrocortisone improves sensitivity of peripheral tissues to insulin, it may be an interesting candidate for use in reducing insulin resistance in peripheral tissues of horses with several disease states. In contrast, long-term administration of eGH decreased tissue sensitivity to exogenous insulin associated with hyperinsulinemia. Therefore, increased concentrations of growth hormone may contribute to insulin resistance in horses with various disease states.

Phenytoin sodium as a treatment for ventricular dysrhythmia in horses.

Five adult horses with ventricular extra systoles (VES) and 2 with ventricular tachycardia (VT) refractory to treatment with rest, anti-inflammatory drugs, lidocaine, or procainamide were treated with phenytoin sodium p.o. q12h. The starting dosage of phenytoin was 20 or 22 mg/kg body weight (BW) q12h, and the maintenance dosage varied from 8 to 17 mg/kg BW q12h. The mean +/- standard deviation therapeutic blood concentration of total phenytoin was 8.8 +/- 2.1 mg/L, and the mean concentration of free phenytoin of 2.5 +/- 0.5 mg/L was relatively constant at a range of 24 to 29% of the total phenytoin concentration. In all horses, both VES and VT were abolished after treatment with phenytoin. On the basis of the results of this clinical study, the authors propose an initial dose of 20 mg/kg BW q12h for the first 3 or 4 dosages, followed by a maintenance dose of 10 to 15 mg/kg BW q12h. Phenytoin plasma concentrations should be monitored during therapy. High plasma concentrations were associated with adverse effects such as recumbency and excitement. In this study, which included a limited number of diverse patients, phenytoin sodium appeared to be an inexpensive and effective treatment for persistent VES or VT in cases where conventional treatment had failed.

Influence of age of horse on results of quantitative electromyographic needle examination of skeletal muscles in Dutch Warmblood horses.

OBJECTIVE: To determine the influence of age on results of quantitative analysis of electromyographic (EMG) needle examination in the subclavian, triceps, and lateral vastus muscles of Dutch Warmblood horses. ANIMALS: 7 healthy young Dutch Warmblood horses (range, 13 to 18 months old), 7 healthy adult Dutch Warmblood horses (range, 4 to 10 years old), and 7 healthy elderly Dutch Warmblood horses (range, 18 to 21 years old). PROCEDURE: An EMG needle examination was performed to evaluate insertional activity, spontaneous activity, and motor unit action potential (MUAP) variables. Although all horses were conscious, young horses were sedated prior to examination. RESULTS: Mean insertional activity in young horses was significantly lower than in elderly horses. Pathologic spontaneous activity was rarely found in young and adult horses but was frequently evident in all muscles in all elderly horses. The MUAP duration and amplitude were significantly lower in all muscles of young horses, compared with values for adult and elderly horses. The MUAP duration and number of phases and turns were significantly lower in adult horses than in elderly horses. Group differences for percentages of polyphasic and complex MUAPs were also found. The 95% confidence intervals for MUAP duration, MUAP amplitude, and number of phases and turns for the subclavian, triceps, and lateral vastus muscles were significantly lower in young horses than in adult or elderly horses. CONCLUSIONS AND CLINICAL RELEVANCE: Age of the horse being examined should be considered when EMG examination is performed.

Electromyographic changes of motor unit activity in horses with induced hypocalcemia and hypomagnesemia.

OBJECTIVE: To determine whether electromyographic abnormalities are evident in skeletal muscles in horses with induced hypocalcemia and hypomagnesemia. ANIMALS: 7 healthy adult Dutch Warmblood horses. PROCEDURES: Electromyographic examination was performed in the lateral vastus, triceps, and subclavian muscles before and after IV infusion of EDTA. An initial dose (mean +/- SD, 564+/-48 ml) of a 10% solution of sodium EDTA was administered IV during a period of 21+/-73 minutes to establish a blood concentration of ionized calcium of approximately 0.5 mmol/L. Average rate of EDTA infusion to maintain ionized calcium at this concentration was 6.6 ml/min. RESULTS: Mean blood concentrations of ionized calcium and magnesium were 1.39+/-0.06 and 0.84+/-0.09 mM, respectively before EDTA infusion; after EDTA infusion, concentrations were 0.48+/-0.05 and 0.44+/-0.20 mM, respectively. This state induced positive waves; fibrillation potentials; doublets, triplets, and multiplets; complex repetitive discharges; and neuromyotonia. Analysis of motor unit action potentials (MUAP) after EDTA infusion revealed an increase in prevalence of polyphasic and complex MUAP in all muscles. CONCLUSIONS AND CLINICAL RELEVANCE: None of the horses had classical signs of hypocalcemia and hypomagnesemia. In contrast, all horses had spontaneous activity in the measured muscles indicative of nerve hyperirritability. Calcium and magnesium deficits appear to have consequences, which may be subclinical, affecting functions of the neuromuscular system. This is of interest for equestrian sports in which hypocalcemia and hypomagnesemia are expected, such as during endurance rides.

Quantitative analysis of motor unit action potentials in the subclavian muscle of healthy horses.

OBJECTIVE: To evaluate the application of analysis of motor unit action potentials (MUAP) in horses and to obtain values of MUAP for the subclavian muscle of horses. ANIMALS: 10 healthy adult Dutch Warmblood horses. PROCEDURE: Electromyographic examination of the subclavian muscle in conscious nonsedated horses was performed to evaluate insertional activity, spontaneous activity, MUAP variables, and recruitment patterns. Muscle and body temperatures were measured at the beginning and end of the procedure. Amplitude, duration, number of phases, and number of changes in direction (ie, turns) for all representative MUAP were analyzed to determine values for this muscle in this group of horses. RESULTS: Mean +/- SD duration of insertional activity was 471.7 +/- 33.45 milliseconds. Mean MUAP amplitude in the examined horses was 379 RV (95% confidence interval [CI], 349 to 410 microV). Mean MUAP duration of the subclavian muscle was 727 milliseconds (95% CI, 6.84 to 7.71 milliseconds). Mean number of phases was 2.9, and mean number of turns was 3.0. Prevalence of polyphasic MUAP defined as MUAP with > 4 phases, was 77%. Number of MUAP that had > 5 turns was 2.4%. Satellite potentials were found in 1.0% of the MUAP CONCLUSIONS AND CLINICAL RELEVANCE: This study revealed that electromyography including MUAP analysis can be performed in horses, and values for the subclavian muscle in healthy adult horses can be obtained. Analysis of MUAP could be a valuable diagnostic tool for use in discriminating between myogenic and neurogenic problems in horses.

Immunohistochemical identification and fiber type specific localization of protein kinase C isoforms in equine skeletal muscle.

OBJECTIVE: To investigate whether protein kinase C (PKC) isoforms are expressed in equine skeletal muscle and determine their distribution in various types of fibers by use of immunofluorescence microscopy. ANIMALS: 5 healthy adult Dutch Warmblood horses. PROCEDURE: In each horse, 2 biopsy specimens were obtained from the vastus lateralis muscle. Cryosections of equine muscle were stained with PKC isoform (alpha, beta1, beta2, delta, epsilon, or zeta)-specific polyclonal antibodies and examined by use of a fluorescence microscope. Homogenized muscle samples were evaluated via western blot analysis. RESULTS: The PKC alpha, beta1, beta2, delta, epsilon, and zeta isoforms were localized within the fibers of equine skeletal muscle. In addition, PKC alpha and beta2 were detected near or in the plasma membrane of muscle cells. For some PKC isoforms, distribution was specific for fiber type. Staining of cell membranes for PKC alpha was observed predominantly in fibers that reacted positively with myosin heavy chain (MHC)-IIa; PKC delta and epsilon staining were more pronounced in MHC-I-positive fibers. In contrast, MHC-I negative fibers contained more PKC zeta than MHC-I-positive fibers. Distribution of PKC beta1 was equal among the different fiber types. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that PKC isoforms are expressed in equine skeletal muscle in a fiber type-specific manner. Therefore, the involvement of PKC isoforms in signal transduction in equine skeletal muscle might be dependent on fiber type.