Determination of red blood cell potassium content in horses with diarrhoea: A practical approach for therapy

The concentration of potassium in the red blood cells in 15 horses with severe diarrhoea was determined. The red blood cell potassium content (RBCK⁺) was low compared with levels obtained in normal horses. The decrease in the RBCK⁺ was used to evaluate the total body potassium deficit. This calculated amount was administered orally, following each RBCK⁺ determination. In those horses which recovered, RBCK⁺ content returned to normal levels. During treatment, refilling of the red blood cells with potassium occurred only gradually as long as the diarrhoea persisted. In each case, oral loading with the calculated dose resulted in an improvement of the general condition and no side effects were observed. The importance of using RBCK⁺ levels as a diagnostic and therapeutic parameter is discussed.     

Thrombocytopenia in the cavalier King Charles spaniel

Blood samples were collected for the determination of platelet counts from 102 cavalier King Charles spaniels which had no history of bleeding disorders. Thrombocytopenia was found in 31 per cent of the samples when the lower reference limit was set at 100 X 10⁹/litre. The platelet count of the males (mean ± SD, 147 X 10⁹/litre ± 15 X 10⁹/litre) was found to be significantly (P<0.05) lower than that of the females (mean ± SD, 202 X 10⁹/litre ± 13 X 10⁹/litre). The platelet counts from a reference group of 16 normal beagles, analysed identically, were not lower than 189 X 10⁹/litre and no significant difference was found between males (mean ± SD, 249 X 10⁹/litre ± 14 X 10⁹/litre) and females (mean ± SD, 282 X 10⁹/litre ± 20 X 10⁹/litre).     

Genetic-diet interactions in the Hyperkalemic Periodic Paralysis syndrome in Quarter Horses fed varying amounts of potassium: I. Potassium and sodium balance, packed cell volume and plasma potassium and sodium concentrations

Six broodmares that were genetically tested to be heterozygous (H/N) and six broodmares that were tested to be homozygous negative (N/N) for Equine Hyperkalemic Periodic Paralysis (HYPP), that were descendents of the same stallion, were used in a replicated 3×3 Latin square experiment to determine the genetic-diet relationships in the HYPP syndrome.¹ The mares were fed rations consisting of 65% pelleted concentrate and 35% Coastal Bermuda grass hay that provided 1.1 (diet A), 1.9 (diet B) and 2.9% (diet C) potassium. The experimental periods were 14 days long, resulting in 28 meals per period. At meals 1 and 27, blood samples were taken every 30 minutes for 12 hours and every 10 minutes from 2 to 5 hours post-feeding. Total urine and fecal collections were made on the last 4 days of each period.Water, feed and fecal samples were analyzed for sodium and potassium content. Blood was analyzed for packed cell volume (PCV) and plasma potassium (K⁺) and sodium (Na⁺) concentrations.Apparent absorption of potassium was 99.8% across all diets, and potassium was excreted principally in the urine. More potassium was retained when the horses were fed diet B than diets A or C. Apparent absorption of sodium was 99.6% with no difference by diet. Neither potassium nor sodium balances were affected by HYPP status.There was no post-prandial pattern of plasma K⁺ concentration seen when the horses were fed the low potassium diet. When fed the higher potassium diets, they had greater plasma K⁺ values and exhibited a post-prandial peak at 2 to 5 hours after feeding. This peak occurred in all of the horses, meaning that plasma K⁺ cannot be used as a diagnostic aid for the presence of the HYPP mutation. By meal 27, some adaptation had occurred resulting in lower plasma K⁺ values when the horses were fed the higher potassium diets. The horses had increased plasma K⁺ concentrations throughout the post-prandial period while fed diet B compared to diet A, agreeing with the positive potassium balance seen in this experiment. The HYPP H/N horses had lower plasma Na⁺ concentrations in the early post-prandial hours and continuously greater PCV values than the HYPP N/N horses.     

Physiological stimuli of thirst and drinking patterns in ponies

The stimuli that elicit thirst were studied in four ponies. Nineteen hours of water deprivation produced an increase in plasma protein from 67 +/- 0.1 g/litre to 72 +/- 2 g/litre, a mean (+/- se) increase in plasma sodium from 139 +/- 3 to 145 +/- 2 mmol/litre and an increase in plasma osmolality from 297 +/- 1 to 306 +/- 2 mosmol/litre. Undeprived ponies drank 1.5 +/- 0.9 kg/30 mins; 19 h deprived ponies drank 10.2 +/- 2.5 kg/30 mins and corrected the deficits in plasma protein, plasma sodium and plasma osmolality as well as compensating for the water they would have drunk during the deprivation period. In order to determine if an increase in plasma osmolality would stimulate thirst, 250 ml of 15 per cent sodium chloride was infused intravenously. The ponies drank when osmolality increased 3 per cent and when plasma sodium rose from 136 +/- 3 mmol/litre to 143 +/- 3 mmol/litre. Ponies infused with 15 per cent sodium chloride drank 2.9 +/- 0.7 kg; those infused with 0.9 per cent sodium chloride drank 0.7 +/- 0.5 kg. In order to determine if a decrease in plasma volume would stimulate thirst, ponies were injected with 1 or 2 mg/kg bodyweight (bwt) frusemide. Plasma protein rose from 68 +/- 2 g/litre pre-injection to 75 +/- 2 g/litre 1 h after 1 mg/kg bwt frusemide and to 81 +/- 1 g/litre 1 h after 2 mg/kg bwt frusemide.(ABSTRACT TRUNCATED AT 250 WORDS)     

Plasma endotoxin levels in horses subjected to carbohydrate induced laminitis

Thirteen (65 per cent) of 20 horses subjected to carbohydrate overload developed Obel Grade 3 lameness within 56 h. Increases in plasma endotoxin from control levels of less than 0.1 ng/litre to values ranging from 2.4 to 81.53 ng/litre were measured in 11 (85 per cent) of 13 horses during the onset of Obel Grade 3 lameness. Obel Grade 3 lameness was associated with rises in plasma Gram-negative endotoxin levels in 11 (92 per cent) of 12 horses. Two peak increases separated by 16 h were verified in five (45 per cent) of 11 horses that exhibited both endotoxaemia and Obel Grade 3 lameness. The first peak occurred, on average, at 32 h and the second peak at 48 h post overload when only one peak was measured, this occurred in four horses at the average time of 24 h whereas in another three horses only a 48 h or 56 h peak was detected after carbohydrate overload.     

Iatrogenic hypocalcaemia during parenteral fluid therapy of diarrhoeic calves

Acid-base balance and electrolyte concentrations, including ionised calcium, were monitored during intravenous fluid therapy of 11 collapsed diarrhoeic suckler calves aged five to 10 days. Six healthy calves of similar age and type were used to provide control data. All the diarrhoeic calves were severely acidotic (TCO2<12 mmol/litre). Isotonic sodium bicarbonate (1-3 per cent) was administered until the metabolic acidosis was half corrected, as indicated by the TCO2 increasing to 17 to 24 mmol/litre when the infusion was changed to an extracellular volume replacement fluid containing 144 mmol/litre Na+, 35 mmol/litre HCO3-, 4 mmol/litre K+ and 113 mmol/litre Cl- which was administered until the calf was discharged. Milk feeding was started as soon as the calf had a suck reflex. The treatment was successful in 10 calves. At admission the diarrhoeic calves were hypocalcaemic compared with the control calves, but their ionised calcium was significantly higher, with significantly less calcium being protein bound. Treatment with isotonic sodium bicarbonate resulted in a significant improvement in acid-base balance, but both total and ionised calcium decreased significantly, the decrease in ionised calcium being proportionately greater owing partly to a significant increase in the protein binding of calcium. The mean total, bound and ionised calcium concentrations were all significantly lower in the treated calves after they had received isotonic sodium bicarbonate than in the control calves. Further treatment with replacement fluid had no significant effect on any of the parameters apart from pCO2 which increased significantly. Milk feeding had no significant effect on plasma calcium concentrations. The calves' mean ionised calcium concentration was significantly lower at the end of the treatment than before it, but there was no difference in the mean total and bound calcium concentrations. The calves' mean plasma potassium and magnesium concentrations decreased significantly during the course of the treatment.     

Evaluation of the basal and post-adrenocorticotrophic hormone serum concentrations of 17-hydroxyprogesterone for the diagnosis of hyperadrenocorticism in dogs

Serum concentrations of 17-hydroxyprogesterone and cortisol were measured before and after the administration of exogenous adrenocorticotrophic hormone (ACTH) to three groups of dogs: 27 healthy dogs (group 1), 19 dogs with non-adrenal illness (group 2) and 46 dogs with hyperadrenocorticism (group 3). The median (range) post-ACTH concentrations of 17-hydroxyprogesterone were 5.0 (22.2 to 16.8), 6.9 (2.0 to 36.2) and 14.4 (1.7 to 71) nmol/litre in groups 1, 2 and 3, respectively. There were no significant differences in the basal or post-ACTH concentrations of cortisol or 17-hydroxyprogesterone between groups 1 and 2. The post-ACTH concentrations of 17-hydroxyprogesterone in group 3 were significantly (P<0.001) greater than those in groups 1 and 2 combined. The area under the receiver operating curve (ROC) for the post-ACTH concentration of cortisol (0.94) was significantly greater than that for the post-ACTH concentration of 17-hydroxyprogesterone (0.76). Using a two-graph ROC analysis, a cut-off of 8.5 nmol/litre was found to maximise both the sensitivity and specificity of the post-ACTH concentration of 17-hydroxyprogesterone for the diagnosis of hyperadrenocorticism at 71 per cent. With a cut-off of 4.5 nmol/litre the sensitivity increased to 90 per cent but the specificity decreased to 40 per cent; with a cut-off of 16.7 nmol/litre the specificity increased to 90 per cent but the sensitivity decreased to 47 per cent.     

Urinary cortisol:creatinine ratios in healthy horses and horses with hyperadrenocorticism and non-adrenal disease

Urinary cortisol and creatinine concentrations, and the cortisol:creatinine ratio were compared between 12 healthy horses (group 1), 13 horses with Cushing's disease (group 2), and eight horses with dysautonomia syndrome (equine grass sickness) (group 3). The mean (sd) urinary cortisol concentrations were 112 (55.7), 250 (357) and 864 (526) nmol/litre in groups 1, 2 and 3, respectively; the mean (sd) urinary creatinine concentrations were 18.9 (7.3), 12.0 (6.7) and 45.2 (26.4) nmol/litre in groups 1, 2 and 3, respectively, and the mean (sd) ratios were 6.1 (2.6), 19.8 (23.8) and 21.3 (14.5) (x 10(-6)) in groups 1, 2 and 3, respectively. The urinary cortisol and creatinine concentrations were significantly greater in group 3 than in groups 1 and 2, but the ratios were not significantly different, although there was a trend (P=0.076) towards higher values in groups 2 and 3. A diagnostic cut-off in the cortisol:creatinine ratio for the confirmation of Cushing's disease of more than 6.9 x 10(-6) was associated with a diagnostic sensitivity and specificity of 92.3 and 75.0 per cent, respectively, when compared with healthy horses. However, when group 3 horses were included, a cut-off of more than 7.4 x 10(-6) was associated with a diagnostic sensitivity and specificity of 84.6 and 54.5 per cent, respectively.     

Correlations between potassium, calcium and magnesium in blood and muscle of standardbred trotters

This study was conducted to investigate the relationships between the concentrations of potassium, sodium, calcium and magnesium of plasma and muscle in horses, as interactions between these ions can be important in the development of muscle cell injury. Blood samples and gluteal muscle biopsies were taken from 20 healthy Standardbreds to determine the potassium concentration in plasma (cK⁺(P)), red blood cells (cK⁺(RBC)), and muscle (cK⁺(M)), and the calcium and magnesium concentrations in plasma and muscle. No correlations were found between cK⁺(P), cK⁺(RBC), and cK⁺(M). Significant positive correlations were found between cK⁺(M) and muscle magnesium (p<0.01); and cK⁺(P) and plasma calcium (p<0.005). Significant negative correlations were found between cK⁺(M) and the percentage of muscle water (p<0.005); and the muscle magnesium and calcium concentrations (p<0.02).     

Some Effects of a Low Sodium Diet High in Potassium on the Renin-Angiotensin System and Plasma Electrolyte Concentrations in Normal Dogs

Pedersen, H.D., J. Koch, A.L. Jensen, K. Poulsen, A. Flagstad: Some effects of a low sodium diet high in potassium on the renin-angiotensin system and plasma electolyte concentrations in normal dogs. Acta vet. scand. 1994, 35, 133-140. - Eight normal male Beagle dogs received 0.7 mmol Na⁺/kg/day for 5 weeks and 4.0 mmol Na⁺/kg/day in one 3 week control period preceding and another similar period following the low sodium period. The dogs received 6.8 mmol K⁺/kg/day throughout the study.The median plasma renin activity (PRA) and plasma aldosterone concentration (PAC) were higher in the low sodium period than in the following control period (0.67 versus 0.28 ng/ml/h, p < 0.0001) and (204 versus 31 pg/ml, p < 0.0001). PRA and PAC quickly stabilized on a new steady level in response to altered intake of sodium chloride. The angiotensin-converting enzyme (ACE) activity was not changed by the altered intake of sodium chloride.The plasma concentrations of sodium and chloride were increased during the low sodium period. This could be due to an indirect effect of the high potassium intake of the dogs. Potassium leads to an increased secretion of aldosterone and thereby to an increased retention of sodium and chloride in the kidney. The possible implications of a high potassium content in a low sodium diet are discussed.     

External balance of water and electrolytes in the horse

The components of the daily external water and electrolyte balances of five normal Standardbred geldings with a mean body weight of 427 kg, were determined over 14 day periods. The mean value of daily total water intake (a summation of free, combined and metabolic water intake) for the group was 27.6 litres, that is, 64.4 ml/kg body weight, while mean values for ambient temperature and humidity ranged from 16 to 25 degrees C and 55.4 to 75.5 per cent, respectively. The mean daily urine volume of 9.9 litres (23.2 ml/kg) for the group, was found to exceed the mean faecal water output of 7.2 litres (16.9 ml/kg). The mean output of sodium for the five animals, in urine and faeces, was 780 mmol or 79 per cent of the intake. The mean output of potassium, chloride, calcium and magnesium were 3163, 2521, 1824 and 1448 mmol or 95, 84, 74 and 75 per cent, respectively of intake. The sodium and potassium not accounted for in urine and faeces may well have been lost in sweat. Results are discussed and compared with those of a previous external balance study in the horse.     

Effect of oral L-carnitine on its concentration in the plasma of yearling Thoroughbred horses

After an oral dose of 10 g of L-carnitine the plasma concentrations of free and total carnitine increased in five yearling thoroughbred horses, reaching a peak two to four hours after administration, but in two horses there was no increase. In the five which responded, the mean (+/- sd) peak increase in total carnitine concentration was 15.1 +/- 6.9 mumol/litre from a predose mean of 18.8 +/- 3.4 mumol/litre, and the mean peak increase in free carnitine concentration was 13.5 +/- 3.1 mumol/litre from a pre-dose mean concentration of 12.5 +/- 2.2 mumol/litre. These changes were similar to those observed previously in a group of adult thoroughbred horses given 10 g of L-carnitine. There was no consistent pattern of change in the concentration of acetylcarnitine in the seven yearling horses.     

Review of 30 cases of peritonitis in the horse

Thirty cases of peritonitis, in which the diagnosis was based on a peritoneal fluid white blood cell count in excess of 10 x 10(9)/litre, are described. Colic, ileus, pyrexia, weight loss and diarrhoea were common presenting signs. Treatments included intravenous fluids, anti-inflammatory analgesics, broad spectrum antibiotics and anthelmintics. Duration of treatment was determined by the clinical condition of the horse and sequential analyses of the peritoneal fluid and the haemogram. In the majority of cases the primary cause of peritonitis was not accurately determined, but 21 horses (70 per cent) recovered. All the horses with diarrhoea were killed after marked deterioration in their clinical condition despite intensive treatment. No individual laboratory parameter was of value in determining prognosis, although of the eight (27 per cent) horses from which bacteria were identified in the initial peritoneal fluid by Gram stain, four (50 per cent) were subsequently killed.     

Radioactive iodine therapy for feline hyperthyroidism: Efficacy and administration routes

The efficacy of radioactive iodine (¹³¹I) administration was studied in a series of 50 hyperthy-roid cats. The dose administered to each cat was based on the clinical severity of the thyrotoxicosis, the serum total thyroxine (TJ concentration and the size of the goitre estimated by palpation. The activity ranged from 80 to 200 MBq (mean ± SD, 143 ± 24 MBq}. The ¹³¹I was injected intravenously in 27 cases and subcutaneously in 23 cases. Each cat was hospitalised for 30 days after the injection. Regardless of the route of injection, none of the cats exhibited any side effects after therapy and all tolerated the hospitalisation period well. There was a significant (P<0.001) decrease in the serum total T₄ concentration (reference range, 10.4 to 42.0 nmol/litre) from a mean ± SD of 181.3 ± 111.4 nmol/litre (range, 43.8 to 575.6 nmol/litre) to a mean ± SD of 19.0 ± 29.6 nmol/litre (range, 2.0 to 175.7 nmol/litre) 30 days following the injection of the radioisotope. Five cats remained hyperthyroid, although in each case the serum total T₄ concentrations had decreased from pre-treatment values. Two of the cats subsequently became euthyroid within three and five months of therapy, respectively, two were lost to adequate follow-up and the remaining cat was successfully retreated four months later. Before treatment, four of these cases had high scores based on the three criteria used for dose estimation. Serum total T₄ concentrations below the reference range developed after treatment in many cases, but were often transient. Clinical evidence of hypothyroidism was not apparent in any cat. Recurrence of hyperthyroidism has not occurred in follow-up periods of up to 32 months. There was no difference in the outcome between the cats injected intravenously or subcutaneously and the latter was considered to be safer and simpler. The administration of an approximated dose of ¹³¹I proved to be an effective treatment for hyperthyroidism in 47 (94.0 per cent) of the cats and obviated the need for sophisticated nuclear computer facilities. There may be a lag period in some cases before euthy-roidism is achieved and this should be considered before the administration of a second dose. ¹³¹I can be administered subcutaneously without untoward effects.     

Hyperkalemic periodic paralysis, plasma lactate and exercise tolerance

The purpose of this, trial was to determine the effect of hyperkalemic periodic paralysis (HYPP) on exercise tolerance in Quarter Horses. Five HYPP affected and five nonaffected horses were matched for age, size, gender and reproductive status. HYPP status was diagnosed by DNA analysis and potassium chloride challenge testing. Plasma lactate concentration and heart rate were used as indicators of work intensity. Serum potassium concentrations were also monitored. Two exercise experiments were conducted, the first being forty-five minutes of slow, aerobic exercise (hacking) and the other being moderate, partially anaerobic exercise (galloping). Post-exercise the horses were cooled out by randomly assigning them to either forty minutes, of standing still or forty minutes of walking. Heart rates of HYPP affected and unaffected horses were not significantly different during exercise or recovery. Plasma lactate concentrations changed slightly following slow exercise and were significantly higher for HYPP affected horses (P=0.01).At the end of exercise, values were 1.4±0.2 mmol/L and 1.0±0.1 mmol/L for HYPP affected and unaffected horses, respectively. Following moderate exercise, plasma lactate concentrations were much greater, and the difference (P<0.001) between affected and unaffected horses was more marked: immediately following exercise concentrations were 10.6±1.8 and 6.2±1.0 mmol/L in affected and unaffected horses, respectively. The higher post-exercise plasma lactate concentrations in affected horses indicates increased anaerobic muscle metabolism. Serum potassium concentrations rose following exercise and significantly higher values were seen in horses that were walked rather than stood still post-exercise.Hyperkalemic periodic paralysis (HYPP) is a dominant autosomal genetic defect occurring in American Quarter Horses and related breeds. ^1–5 The condition is widely geographically distributed and has been estimated to affect 0.4% of all Quarter Horses.⁶ Signs include sporadic attacks of muscle fasciculation, muscle spasm, sweating and weakness. Oral administration of potassium chloride produces a more severe hyperkalemia in HYPP affected horses and induces clinical attacks.¹ Electromyography reveals widespread continuous, spontaneous, muscle contraction.² Some people believe that HYPP affected horses are suitable for riding⁷; others have expressed doubts about the safety of this practice and the exercise tolerance of affected horses. HYPP affected horses have an unstable muscle membrane potential causing random muscle fiber contractions,^2,8 which could antagonize purposeful movement. This, in combination with increases in extracellular potassium concentration occurring during exercise,^9–11 may cause affected horses to stumble or collapse while being ridden, posing a danger to both horse and rider. In HYPP affected people, potassium concentration rises approximately one to two hours after exercise during which time HYPP attacks can occur.^9,10 For these reasons, the exercise tolerance of HYPP affected horses and the effects of exercise on spontaneous HYPP attacks deserves investigation.We standardized the exercise test and chose heart rate and plasma lactate concentrations as indicatoors, of exercise tolerance and energy metabolism.¹² Heart rate is the major determinant of oxygen delivery to muscle and the rate rises with exercise intensity until it reaches a plateau at high velocities.^13,14 Several lines of investigation indicate that plasma lactate concentrations reflect muscle work. Lactate release from resting muscle is minimal but large amounts are released during strenuous exercise.^15–17 Lactate release increases with increasing muscle work.^16,18 During exercise, muscle oxygen consumption increases until eventually a plateau is reached after which no further increase in oxygen consumption in response to increased work loads is possible.¹⁸ At this point the mitochondrial electron chain transport system is operating at its maximal possible rate, energy production can only be supplemented by the use of inefficient anerobic glycolytic metabolism and the muscle starts to release lactate. If the concentrations of plasma lactate are high, the muscles are depending on anaerobic metabolism and are working closer to exhaustion^16,18–21The objectives of this experiment were twofold. One was to determine if HYPP adversely affects exercise performance. The second was to study the effects of cooling out, by either standing still or walking, on serum potassium and plasma lactate concentrations.     

Field study of the prevalence of lameness in horses with back problems

A population of 805 horses (70 per cent dressage, 20 per cent show jumpers and 10 per cent trotters) with orthopaedic problems was examined for signs of lameness and back problems, irrespective of their original complaints. In the horses with a back problem the prevalence of lameness was 74 per cent, and back problems were diagnosed in 32 per cent of the lame horses. These percentages were significantly higher than those recorded in a control population of 399 horses, of which 20 per cent were lame and 12 per cent had back problems. In the group of horses with orthopaedic problems there was a strong association between lameness and back problems and, in particular, there was a high prevalence of lameness among the horses with back problems.     

Mammary secretions in normal spontaneous and induced premature parturition in the mare

Total calcium, total protein, albumin and globulin content were determined in 49 samples of mammary secretions from 37 crossbred and Thoroughbred mares. The mares were divided into three categories: Group 1--post partum samples from spontaneous full term Thoroughbred deliveries (n = 20); Group 2--pre- and post partum samples from spontaneous full term Thoroughbred deliveries (n = 6); Group 3--pre- and post partum samples from induced premature deliveries in crossbred mares (n = 11). Calcium concentrations of the mammary secretions proved useful in predicting full term and also in assessment of the chances of foal survival in prematurely induced parturition. Mean (+/- se) calcium levels in full term mares that foaled spontaneously (Group 1) were 10.6 +/- 2.7 mmol/litre on the day of parturition. However lower values than this did not necessarily mean parturition would not take place. In Group 2 mares, calcium levels were 4.1 +/- 0.8 mmol/litre just before foaling and rose to 9.2 +/- 1.5 mmol/litre within 20 mins of parturition; an increase of 124 per cent. Group 3 mares, induced prematurely, showed poor foal survival if pre-partum calcium levels were less than 3 mmol/litre. Total protein, albumin and globulin concentrations showed a similar trend but the increase just before or during parturition was much less marked.     

Detection of bicarbonate administration (milkshake) in Standardbred horses

SUMMARY Total plasma carbon dioxide (TCO₂) concentrations were measured in Standardbred horses to determine criteria to discriminate between normal horses and horses with excessive TCO₂ concentrations on raceday. TCO₂ concentrations from stabled horses were distributed normally with a mean of 30.2 mmol/L and a standard deviation of 1.2 (n = 192) while pre-race TCO₂ concentrations were not normally distributed. The results indicate that about 50 horses per million are likely to have TCO₂ concentrations greater than or equal to 35 mmol/L and that it is extremely unlikely that a normal horse would have a resting TCO₂ concentration above 36 mmol/L. These values were associated with sensitivities of 67% and 59%, respectively, and with a specificity of 100%. TCO₂ concentrations were relatively stable in blood samples stored at 4°C for 4 days, whereas the TCO₂ in specimens stored at room temperature (25°C) and at ambient temperature (16–28°C) declined progressively over 5 days. The accuracy and precision of the Beckman EL-ISE Auto Analyser were acceptable and within the manufacturers specified range. Paired specimens analysed using a Beckman EL-ISE Auto Analyser and a Kodak Dry Chemistry Analyser were not significantly different. However, the measurements made using the Kodak Dry Chemistry Analyser averaged 0.5 mmol/L higher than those analysed on the Beckman EL-ISE. The significance of these sources of variation in TCO₂ concentration in relation to the testing of horses for ‘milkshake’ administration are discussed.     

An automated micro-method for the enzymatic determination of D-B-hydroxybutyrate in ruminant plasma

1. An enzymatic ‘reaction rate’ micro-method for the rapid routine estimation of D-B-hydroxybutyrate (D-B-HOB) in ruminant plasma, using an I.L. Multistat III centrifugal analyzer, is described.
2. Reaction conditions were optimized to give a linear response for plasma D-B-HOB concentrations between 100 and 2500 μmoles per litre, at 30°C and pH 9.0.
3. For the standardized method the within-run and between-run coefficients of variation for deproteinised ovine plasma were consistently less than 3.5%.
4. There was good agreement between plasma concentrations obtained by the present method and both original U.V. end-point technique (r=0.927b=0.950) and a colorimetric end-point procedure (r=0.937. b=0.879).
5. Utreated ovine and bovine plasma consistently exhibited high ‘blank’ activity and this was directly correlated with plasma lactate dehydrogenase (LDH) activity in both species (r=0.971; p<0.001 and r=0.949; p<0.001 respectively). The distribution of LDH activity in man was similar to sheep but, contrastingly, non-specific interference was extremely low in human plasma and unrelated to LDH. Horse, chicken and rat had negligible ‘blank” activity and comparatively low LDH levels. In both cattle and sheep non-specific interference was abolished by perchloric acid precipitation. In the sheep subtraction of ‘blank’ activity gave D-H-HOB concentrations for untreated plasma comparable to those in deproteinised samples. However, in the bovine, D-B-HOB levels remained significantly (t=6.44; p<0.001) higher even after ‘blank’ correlation. In contrast to man and other non-ruminants, perchloric acid precipitation is essential in ruminants to avoid false overestimation of plasma D-B-HOB levels.
6. Plasma with EDTA as anticoagulant and serum gave concentrations of D-B-HOB approximately 60% lower, than samples containing heparin or oxalate/fluoride. However, heparin was associated with much higher (up to 50%) non-specific NAD rduction than oxalate/fluoride.
7. High levels of acetoacetate (400–1000 μmoles per litre) reduced the recovery of D-B-HOB from ovine plasma by less than 10%. This effect was negated by the inclusion of hydrazine hydrates in the reaction mixture. Perchlorate ion concentrations above 25 μmoles per litre per test dramatically inhibited the assay in ovine plasma, and therefore precipitation conditions must be carefully controlled.
8. Plasma with oxalate/fluoride as anticoagulant showed the greatest stability in storage; 24 hours at room temperature, one week at +4°C and at least one month at ?20°C.

     

Postmortem biochemical markers of experimentally induced hypomagnesaemic tetany in sheep

Hypomagnesaemic tetany was induced in non-lactating and lactating ewes by feeding them semi-synthetic low magnesium diets containing additional potassium chloride and citric acid. Aqueous and vitreous humour were sampled from one eye at the time of death (fresh) and from the second eye after the head had been stored at ambient temperature for 24 hours (24-hour). There were significant relationships between the concentrations of magnesium in cerebrospinal fluid and plasma and its concentrations in fresh aqueous humour and fresh vitreous humour. Magnesium concentrations of < 0.33 mmol/litre in fresh aqueous humour and < 0.50 mmol/litre in 24-hour aqueous humour were associated with severe hypomagnesaemia and tetany. However, the concentration of magnesium in aqueous humour is relatively unstable and, unless the time of death was known accurately, its interpretation would be difficult. Magnesium concentrations of < 0.60 mmol/litre in fresh vitreous humour and < 0.65 mmol/litre in 24-hour vitreous humour were associated with severe hypomagnesaemia and tetany in adult sheep. The concentration of magnesium in vitreous humour was relatively stable for up to 48 hours postmortem.