Palmar digital vessel relaxation in healthy horses and in horses given carbohydrate

OBJECTIVE: To compare in vitro smooth muscle relaxation of palmar digital vessels from healthy horses with those from horses in the prodromal stage of experimentally (carbohydrate) induced laminitis. ANIMALS: 16 adult horses. PROCEDURE: Segments of palmar digital vessels were obtained from 5 healthy horses and 6 horses given carbohydrate. Vascular rings from the palmar digital artery and vein were suspended in individual organ baths containing buffer solution and indomethacin; isometric tension was recorded, and contraction and relaxation were compared. Smooth muscle contraction in response to cumulative addition of phenylephrine was recorded in the absence and presence of 1 microM NG-nitro-L-arginine methyl ester (L -NAME). After wash out, vascular rings were preconstricted with phenylephrine (0.3 microM), and cumulative endothelium-dependent (acetylcholine-induced) and independent (nitroprusside-induced) smooth muscle relaxations were recorded in the absence or presence of L -NAME. RESULTS: Phenylephrine increased vascular smooth muscle tone in ring preparations of palmar digital arteries and veins. Addition of acetylcholine or nitroprusside induced relaxation of palmar digital artery and vein ring preparations. Use of L-NAME (1 microM) significantly reduced maximal relaxation induced by acetylcholine, but not by nitroprusside. Maximal relaxation induced by acetylcholine, but not by nitroprusside, was reduced in vascular rings prepared from carbohydrate-overloaded horses. CONCLUSION AND CLINICAL RELEVANCE: Reduced endothelium-dependent relaxation of palmar digital vessels may have a role in the pathophysiology of acute laminitis after carbohydrate overload in horses.     

Characterization and comparison of the responses of equine digital arteries and veins to endothelin-1

OBJECTIVE: To compare the responses of equine digital arteries (EDAs) and equine digital veins (EDVs) to endothelin-1 (ET-1) and determine the role of the endothelium and type of receptors involved in the modulation and mediation of those responses, respectively. SAMPLE POPULATION: 5 to 9 palmar digital vessels/experiment from 28 healthy horses. PROCEDURE: Rings of dissected vessels were mounted under tension between force transducer wires in organ baths containing Krebs-Henseleit solution at 30 degrees C. Responses of EDAs and EDVs (with intact [+e] or denuded [-e] endothelium) to cumulative concentrations of ET-1 (10(-10) to 3 X 10(-7) M) were compared. For (+e)EDAs and (+e)EDVs precontracted with a thromboxane-mimetic (U44069; 10(-8) M) and (-e)EDAs and (-e)EDVs, responses to an ETB receptor agonist (S6c; 10(-10) to 3 X 10(-7) M) were evaluated. Responses to ET-1 (10(-7) M) in (-e)EDAs and (-e)EDVs were evaluated after incubation with an ETA receptor antagonist (BQ-123; 3 X 10(-7) M), an ETB receptor antagonist (BQ-788; 3 X 10(-7) M), or vehicle solution. RESULTS: Endothelin-1 induced a concentration-dependent contraction of endothelium-intact and -denuded EDAs and EDVs; EDVs were more sensitive. Neither vessel type relaxed in response to S6c, although 2 of the (-e)EDAs contracted mildly. Whereas BQ-123 inhibited the (-e)EDA and (-e)EDV responses to ET-1, BQ-788 had no effect. CONCLUSIONS AND CLINICAL RELEVANCE: Endothelin-1 induced digital vasoconstriction (marked constriction in veins). This action was unaffected by endothelium and mediated predominantly by ETA receptors. These findings suggest ET-1 can induce selective digital venoconstriction.     

Pharmacological characterization of alpha1-adrenoceptors in equine digital veins

Alpha-adrenoceptors mediate contractile responses in equine digital veins (EDVs) and arteries. Vascular smooth muscle alpha(1)-adrenoceptor subtypes have been implicated in a number of conditions, such as acute equine laminitis, and are therapeutic targets for the treatment of this condition. Digital veins, rather than arteries, were investigated in the present study because they have been specifically implicated in the pathophysiology of acute laminitis. The order of potency of a series of alpha(1)-adrenoceptor-selective agonists and antagonists was determined in isolated rings of EDVs under conditions of isometric tension. A61603 was the most potent agonist, with a higher potency (76-fold greater) than phenylephrine (PHE), suggesting the presence of the alpha(1A)-adrenoceptor subtype. Prazosin (30 nm) caused competitive inhibition of the responses to A61603 and PHE, with pK(b) values of 8.05 +/- 0.28 and 8.20 +/- 0.27, respectively. In addition, the alpha(1A)-adrenoceptor antagonist, WB4101 (10 nm), also caused competitive inhibition of the responses to the two agonists, with pK(b) values of 8.37 +/- 0.16 and 8.54 +/- 0.23, respectively, confirming the presence of the alpha(1A)-adrenoceptor subtype in EDVs. The selective alpha(1D)-adrenoceptor antagonist, BMY7378 (100 nm) did not cause a significant change in the response to the agonists, giving lower pK(b) values (6.97 +/- 0.27 and 6.88 +/- 0.17 vs. A61603 and PHE, respectively). Chloroethylclonidine dihydrochloride (45 microm, 30 min), used to produce selective inactivation of alpha(1B)-adrenoceptors, caused noncompetitive inhibition of the response to PHE, but was without effect on the response to A61603. These findings indicate that EDVs possess at least two different alpha(1)-adrenoceptor populations, which are predominantly of the alpha(1A) and alpha(1B) subtypes. These data may assist in the development of more selective antagonists for therapeutic use in horses.     

α- and β-adrenoceptors in the sheep urinary bladder

A study was undertaken to determine the presence and distribution of alpha- and beta-adrenoceptors in the sheep bladder body and base. In the bladder body, noradrenaline and isoproterenol induced relaxation which was significantly inhibited by propranolol, pafenolol and butoxamine. In the presence of propranolol (10(-5) M), noradrenaline induced a small contraction, as well as phenylephrine, but B-HT 920 failed to cause any effect on the bladder body. In the bladder base, noradrenaline caused a contraction that was significantly inhibited by prazosin but not by yohimbine. Phenylephrine also induced a contractile response in this structure which was inhibited by prazosin. Isoproterenol caused a relaxation that was significantly inhibited by propranolol and pafenolol but not by butoxamine. Relaxation was mediated by both beta 1 and beta 2-adrenoceptors in the detrusor muscle and by beta 1-adrenoceptors in the bladder base. Alpha 1-adrenoceptors contributed to maintain the detrusor tone and contract the bladder base.     

Lymphoscintigraphy of draught horses with chronic progressive lymphoedema

REASONS FOR PERFORMING STUDY: Early diagnosis of chronic progressive lymphoedema (CPL) may result in more effective interventions and provide a basis for further investigation of whether early diagnosis could be used as a means of eliminating potential genetic influences by cessation of breeding from affected individuals. HYPOTHESIS: Lymphoscintigraphy may be useful in draught horses to differentiate early lesions of CPL from other conditions in the pastern region. METHODS: Forelimbs of 2 normal and 5 CPL-affected draught horses were evaluated with lymphoscintigraphy. RESULTS: Lymphoscintigraphy showed clearly the presence of interstitial fluid stasis and delayed lymphatic drainage in the affected extremities of diseased animals in contrast to normal animals of these breeds. The rate of decreased clearance of a particulate radiopharmaceutical from the tissues was related positively to the severity of clinical signs. CONCLUSIONS AND POTENTIAL RELEVANCE: Our findings support the hypothesis that lymph stasis is probably responsible for the progressive swelling and concurrent skin lesions observed in association with CPL in draught horses. Lymphoscintigraphy should also prove useful in diagnosis of CPL in draught horses, even in the mild stages of the disease; such early diagnosis may result in more effective intervention.     

Early laminar events involving endothelial activation in horses with black walnut- induced laminitis

OBJECTIVE: To determine proinflammatory gene expression, endothelial adhesion molecule gene expression, and matrix metalloproteinase (MMP) concentrations in laminar specimens at 1.5 hours after administration of black walnut extract (BWE) and to compare these values with later time points. ANIMALS: 25 horses. PROCEDURES: After nasogastric administration of BWE, anesthesia was induced at 1.5 hours in early time point (ETP) horses (n = 5), between 3 and 4 hours in developmental time point horses (5), and between 9 and 10 hours in acute onset of lameness time point horses (5). Anesthesia was induced at 3 and 10 hours after nasogastric administration of water in 2 groups of control horses (3-hour control group, n = 5; 10-hour control group, 5). Real-time quantitative PCR assay was performed on laminar specimens from control and ETP horses for cyclooxygenase (COX)-1, COX-2, interleukin (IL)-1beta, tumor necrosis factor-alpha, IL-6, IL-8, IL-10, MMP-2, and MMP-9 gene expression; and on laminar specimens from all groups for endothelial adhesion molecules, intercellular adhesion molecule (ICAM)-1, and E-selectin gene expression. Leukocyte emigration was assessed via CD13 immunohistochemistry, and gelatinase accumulation was determined by gelatin zymography. RESULTS: Laminar concentrations of IL-1beta, IL-6, IL-8, COX-2, ICAM-1, and E-selectin mRNA were significantly increased in ETP horses, compared with control horses. Concentrations of IL-1beta, IL-8, ICAM-1, and E-selectin mRNA peaked at 1.5 hours. In ETP horses, leukocyte emigration was present in 3 of 5 horses and pro-MMP-9 was detected in 2 of 5 horses. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that endothelial activation and laminar inflammation are early events in laminitis; MMP accumulation likely is a downstream event.     

Characterization of alpha-adrenoceptor subtypes in smooth muscle of equine ileum

OBJECTIVE: To determine the concentration and binding characteristics of alpha-adrenoceptor subtypes in smooth muscle cell membranes of equine ileum. SAMPLE POPULATION: Segments of longitudinal and circular smooth muscle from the ileum of 8 male and 8 female adult horses. PROCEDURE: Distribution of alpha-adrenoceptor subtypes was assessed by use of radioligand binding assays incorporating [3H]-prazosin and [3H]-rauwolscine, highly selective alpha1- and alpha2-adrenoceptor antagonists, respectively. Characterization of adrenoceptor subtypes was performed by use of binding inhibition assays. RESULTS: On the basis of binding affinity for specific radioligands, low- and high-affinity alpha1- and alpha2-adrenoceptors were detected. Concentration of low-affinity alpha2-adrenoceptors was significantly greater in male horses, compared with females. Competition studies confirmed the specificity of the radioligands used in the binding assays. Alpha1-adrenoceptors of both subtypes in male and female horses had a higher affinity for prazosin than phentolamine, whereas yohimbine did not compete with the radioligand for binding. For alpha2-adrenoceptors regardless of subtype, potency of inhibition elicited by each drug varied between sexes. In males, yohimbine was a more potent inhibitor than phentolamine, which was more potent than prazosin. In females, yohimbine was more potent than prazosin, which was more potent than phentolamine. CONCLUSIONS AND CLINICAL RELEVANCE: High- and low-affinity alpha1- and alpha2-adrenoceptors were detected in smooth muscle of equine ileum. Because alpha-adrenoceptor subtypes, particularly alpha2-adrenoceptors, are involved in the regulation of gastrointestinal tract function, characterization of these receptors may represent the basis for development of new therapeutic strategies for the control of gastrointestinal disturbances in horses.     

Effects induced by exercise on lymphocyte β-adrenergic receptors and plasma catecholamine levels in performance horses

The effect of dynamic exercise on complete blood cell count, lymphocyte β-adrenergic receptor and plasma catecholamine (adrenaline and noradrenaline) levels in horses performing different disciplines were investigated during rest and after exercise. Blood samples were collected from jumping horses (n=6), Arabian Endurance horses (n=6) and Standardbred trotters (n=6) before and immediately after competition. Dynamic exercise caused a significant increase in red blood cell count (Standardbred trotters: P=0.0012), haemoglobin concentration (jumping horses: P=0.001; Standardbred trotters: P=0.01), haematocrit percentage (Standardbred trotters: P=0.005), neutrophil percentage (jumping horses: P=0.0003), lymphocyte percentage (jumping horses: P=0.0003), monocyte percentage (Standardbred trotters: P=0.0008), lymphocyte β-AR numbers (jumping horses: P=0.01; Arabian Endurance horses: P=0.016; Standardbred trotters: P=0.05), plasma adrenaline concentration (Standardbred trotters: P=0.0001) and plasma noradrenaline levels (Standardbred trotters: P=0.003). It is concluded that acute increases in plasma catecholamine concentrations depended on the exercise performed and may induce up-regulation of β-AR in equine lymphocytes. However, the exact mechanism of β-AR up-regulation still remains unclear.     

Participation of H1-receptors in histamine-induced contraction and relaxation of horse coronary artery in vitro.

The mechanisms of histamine-induced contraction and relaxation were investigated in rings isolated from a middle part of the left descending coronary arteries of horses. Intact and endothelium-denuded preparations were compared. Rings of horse coronary arteries contracted in response to histamine in a concentration dependent manner, but some of them relaxed with lower concentrations and contracted with higher concentrations. Removal of the endothelium abolished the relaxation and potentiated the contraction. The pD2 values were 4.70 +/- 0.08 in the rings with intact endothelium and 4.95 +/- 0.08 in endothelium-denuded rings. Histamine-induced contractions in intact and denuded preparations were not affected by an H2-antagonist, cimetidine, but were inhibited by an H1-antagonist, diphenhydramine in non-competitive manner in the rings with endothelium and in competitive manner in denuded rings. After precontraction with PGF2 alpha or norepinephrine, histamine relaxed preparations with intact endothelium (pD2 value, 7.80 +/- 0.11), although histamine-induced relaxations were not observed in denuded preparations. The relaxation was competitively inhibited by diphenhydramine. Relaxing response was significantly attenuated by methylene blue, quinacrine, L-nitro-arginine, gossypol and AA861 but not by indomethacin. These results suggest that the histamine-induced contraction and relaxation in horse coronary arteries are mediated mainly by H1-receptors in the smooth muscle and endothelium, respectively, and H1-receptor activation of endothelial cells may liberate vasodilator substances.     

Relaxation of equine tracheal muscle in vitro by different adrenoceptor drugs

Strips of tracheal smooth muscle from 12 horses were contracted by carbachol in tissue baths under isometric conditions. This contraction (≅50% of maximum: EC₅₀) was relaxed completely with adrenoceptor drugs. The only exception was clenbuterol, where the degree of relaxation was ≅90%. In all horses the EC₅₀-value for isoprenaline (mean 1.6 × 10⁻⁸M) was less than that for adrenaline (mean 9.6 × 10^{− 8}M) and noradrenaline (mean 1. 8 × 10_{- 6}M). The potency ratio was 1 < 6 < 110 which indicates that the β₂-subtype dominates among the β-adrenoceptors of equine airways. All preparations were also very sensitive to the specific and potent β₂-receptor agonists clenbuterol (mean 5.7 × 10^{− 9}M) and procaterol (mean 3.6 × 10⁻¹⁰M). No differences in EC₅₀-values due to age, sex and breed were observed in this material. The standard deviation of the mean EC₅₀-values seems to be larger for the specific β₂-adrenoceptor agonists than for the unspecific. A reason for this could be differences in the pattern of the β-adrenoceptor population.     

Ground reaction forces and limb function in tölting Icelandic horses

REASONS FOR PERFORMING STUDY: Gaited horses employ 4-beat stepping (singlefoot) gaits that extend into speeds typical of trots. Ground reaction force (GRF) patterns of these specialised gaits have not been reported; therefore, appraisal of these gaits using nongaited horse kinetics may lead to clinical misjudgements. HYPOTHESIS: GRFs of t?lting Icelandic horses will be comparable in profile and magnitude with those of trotting horses. METHODS: Forelimb and hindlimb GRFs were obtained for 10 Icelandic horses ridden at a t?lt. These data were evaluated across 3 speed ranges: <2, 2.5-5 and >5 m/sec. RESULTS: Virtually all vertical force tracings were single-peaked. Forelimbs typically had greater peak vertical forces and impulses compared with hindlimbs. Support duration and forelimb vertical impulse were correlated negatively with speed, whereas peak vertical, braking and propulsive forces and hindlimb braking and propulsive impulses were correlated positively with speed. CONCLUSIONS: GRF profiles of t?lting Icelandic horses are more similar to profiles of trots than walks, suggesting that t?lts follow bouncing mechanics. POTENTIAL RELEVANCE: Greater overlap of limb support in 4-beat gaits (even at high speeds) is associated with lower peak vertical force magnitudes of t?lts compared with those reported for trots at comparable speeds, which may help limit the occurrence of overloading injuries in Icelandic horses.     

Investigation of mRNA expression of tumor necrosis factor-alpha, interleukin-1beta, and cyclooxygenase-2 in cultured equine digital artery smooth muscle cells after exposure to endotoxin.

OBJECTIVE: To determine messenger RNA expression of cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-alpha, and interleukin- (IL)-1beta from cultured equine smooth muscle cells (SMC). SAMPLE POPULATION: Segments of palmar digital artery harvested from 6 clinically normal adult horses. PROCEDURE: Explants were collected from the tunica media of arteries for primary culture of SMC. Equine mononuclear cells were used as control cells. Subcultured vascular SMC and control cells were exposed to lipopolysaccharide (20 microg/ml and 100 ng/ml, respectively). Northern blot analysis with equine-specific probes for COX-2, TNF-alpha, and IL-1beta was performed, using isolated total cellular RNA. RESULTS: Although no message was detected for IL-1beta or TNF-alpha in control or endotoxin-exposed equine vascular SMC from all horses, COX-2 underwent a distinct substantial up-regulation after endotoxin exposure. Endotoxin-exposed equine mononuclear cells had up-regulation of IL-1beta and TNF-alpha mRNA. CONCLUSIONS AND CLINICAL RELEVANCE: Increased expression of COX-2 mRNA by equine vascular SMC may be an important early pathophysiologic event in the onset of endotoxemia in horses. Potentiated local vascular production of various prostanoids after increased expression of mRNA for COX-2 may result in vasoactive events observed with laminitis.     

Characterization of porcine beta1- and beta2-adrenergic receptors in heart, skeletal muscle, and adipose tissue, and the identification of an atypical beta-adrenergic binding site

The objective of this study was to characterize porcine beta1- and beta2-adrenergic receptors (beta1-AR and beta2-AR) in heart, skeletal muscle, and adipose tissue by measuring the binding of a radioligand to cell membrane fragments. In skeletal muscle (LM), [3H]CGP12177 labeled a homogeneous population of beta2-AR as evidenced by the rank order of affinity of catecholamines [(-)isoproterenol > (-)epinephrine > (-)norepinephrine], a high affinity of the binding site for the beta2-AR-agonist clenbuterol (equilibrium dissociation constant, Kd = 16 nM), and a low affinity of the binding site for the beta1-AR-antagonist CGP20712A (Kd = 21 microM). The affinity of ICI118551, a ligand selective for beta2-AR in other species, was uncharacteristically low in porcine LM (Kd = 441 nM), but was consistent with a value reported for the cloned porcine beta2-AR. In heart ventricle, ligand binding revealed a predominant population of beta1-AR, judged by the rank order of affinity of catecholamines [(-)isoproterenol > (-)epinephrine > or = (-)norepinephrine] and high-affinity binding to CGP20712A (Kd = 40 nM). The Kd for ICI118551 (731 nM) was close to that observed at beta2-AR in LM, confirming that ICI118551 is not subtype-selective in the pig. Displacement studies using (-)propranolol, clenbuterol, and (-)isoproterenol revealed a second high-affinity binding site in the heart that was not a beta2-AR and could not be eliminated by guanosine 5'-triphosphate or guanylyli-midodiphosphate. In adipose tissue, an equal number of beta1- and beta2-AR was identified through the binding of clenbuterol and CGP20712A, whereas ICI118551 could not discriminate between these sites. In further experiments, we used 10 microM CGP20712A to eliminate beta1-AR binding and allow accurate Kd values to be determined at beta2-AR for nonselective ligands. Under these conditions, another binding site was observed that had a high affinity for (-)propranolol (Kd = 20 pM), which is inconsistent with beta3- or beta4-AR binding reported elsewhere. Our results indicate that porcine adipose tissue contains beta1-AR, beta2-AR, and an atypical binding site in the proportions 50, 34, and 16%, respectively, of the total binding sites labeled by [3H]CGP12177.     

Mepartricin long-term administration regulates steroid hormone and adrenergic receptor concentrations in the prostate of aged rats

Mepartricin is a semi-synthetic macrolide antibiotic developed as a drug for the treatment of benign prostatic hyperplasia (BPH) in human patients. In the present study, aged rats are used as an experimental model to evaluate the effects of mepartricin on circulating hormone concentrations and prostate receptor concentrations, to compare these possible effects with clinical findings observed in long-term treated dogs. Fifty-six aged male rats were randomly divided into four experimental groups treated orally with 0 (group 1), 2 mg (group 2), 5 mg (group 3) and 20 mg (group 4) mepartricin/kg of body weight. for 28 days respectively. Serum oestradiol and testosterone concentrations were measured by radio-immune-assays methods. Binding assays were used to measure the prostate concentrations of oestrogen receptors (ER), androgen receptors (AnR), alpha(1)-adrenergic receptor (alpha(1)-AR), and beta-adrenerergic receptor (beta-AR) subtypes. Mepartricin induced a significant reduction of prostate weight and serum oestradiol concentrations. Serum testosterone concentrations were unaffected. The treatment induced a significant down-regulation of ER concentrations (P < 0.05) and a significant up-regulation of AnR (P < 0.05) in rat prostate. Mepartricin induced a significant (P < 0.05) dose-dependent up-regulation of alpha(1)-AR and beta(2)-AR. In contrast, the concentration of beta(3)-ARs was significantly decreased (P < 0.05) in treated animals. The increase in prostate beta(2)-AR concentrations observed in subjects treated with mepartricin may be a favourable element in the evolution of BPH, because of the role exerted by these receptors in the control of prostatic smooth muscle relaxation. Curiously, beta(3)-AR concentrations were significantly reduced in treated animals. Data collected suggest that the prostatic beta-AR expression might be strongly influenced by oestrogen deprivation (mepartricin treatment); therefore, the combination of oestrogen suppression (mepartricin) and adrenergic suppression (alpha(1)-AR blockers) may be proposed as a possible nonhormonal therapeutic strategy for the treatment of benign prostatic hyperplasia in dogs.     

Regulation of equine lymphocyte beta-adrenoceptors under the influence of clenbuterol and dexamethasone

In 12 healthy horses, the effects of the beta2-agonist clenbuterol and the glucocorticoid dexamethasone on the lymphocyte beta2-adrenoceptor density and affinity (determined by (-)-[125I]-iodocyanopindolol binding) as well as its responsiveness (assessed by lymphocyte cyclic AMP [cAMP] responses to 10 micromol/l (-)-isoprenaline) were studied. Clenbuterol treatment, 2 x 0.8 microg/kg/day i.v. for 12 days, decreased significantly ICYP binding sites by approximately 30-40%; concomitantly, lymphocyte cAMP response to (-)-isoprenaline was reduced. After withdrawal of clenbuterol, beta2-adrenoceptor density and responsiveness gradually increased, reaching predrug levels after 4 days. The effects of dexamethasone on clenbuterol-induced desensitisation were further investigated. Administration of dexamethasone (1 x 0.1 mg/kg/day, i.v. for 5 days) immediately after clenbuterol withdrawal accelerated beta2-adrenoceptor recovery: only 24 h after administration dexamethasone restored the number of binding sites and cAMP response to (-)-isoprenaline to levels statistically indistinguishable from values before clenbuterol treatment. Three days after dexamethasone administration, lymphocyte beta2-adrenoceptors were further increased about 2-fold the pretreatment values, and this increase declined gradually after dexamethasone withdrawal, reaching baseline values after 4 days. Furthermore, in groups exposed simultaneously to both drugs, dexamethasone completely prevented clenbuterol-induced decrease in lymphocyte beta2-adrenergic receptor density and responsiveness. No significant change was observed in the dissociation constant for ICYP in any of the situations. We conclude that dexamethasone (glucocorticoids) can reverse and prevent Clenbuterol-induced desensitisation (down-regulation) of the lymphocyte beta2-adrenoceptors and therefore, a combined therapy with clenbuterol and dexamethasone may be potentially beneficial in horses suffering from chronic obstructive pulmonary disease (COPD).     

Response to ractopamine-hydrogen chloride is similar in yearling steers across days on feed

Yearling steers (n = 2,552; 314 kg of initial BW) were used to evaluate the effects of ractopamine-HCl (RAC) and days on feed on performance, carcass characteristics, and skeletal muscle gene expression in finishing steers. Treatment groups included serial slaughter dates of 150, 171, or 192 d on feed. Within each slaughter date, steers either received RAC (200 mg/steer) daily for the final 28 d or were not fed RAC. All steers were initially implanted with Revalor-IS and were reimplanted with Revalor-S after 75 d on feed. At slaughter, muscle samples from the semimembranosus were collected for mRNA analysis of the beta-adrenergic receptors (beta-AR). Ractopamine administration increased (P < 0.05) ADG, G:F, and HCW and increased (P = 0.08) LM area. Ractopamine did not affect the dressing percentage, USDA yield grade, or quality grade (P > 0.3). There was no change in overall feed intake across the entire feeding period; however, feed intake was increased during the 28-d period during which the steers were fed RAC (P < or = 0.05). Greater days on feed decreased (P < 0.05) ADG, G:F, DMI, and the number of yield grade 1 and 2 carcasses. Also, greater days on feed increased (P < 0.05) HCW, dressing percentage, and the number of prime and choice carcasses, as well as the number of yield grade 4 and 5 carcasses. Increasing days on feed decreased (P < 0.05) the abundance of beta(1)-AR and beta(3)-AR mRNA and increased (P < 0.05) the abundance of beta(2)-AR mRNA in skeletal muscle samples obtained at slaughter. Ractopamine had no effect (P > 0.10) on the abundance of beta(1)-AR or beta(3)-AR mRNA, but tended (P = 0.09) to increase beta(2)-AR mRNA. Additional time-course studies with primary muscle cell cultures revealed that advancing time in culture increased (P < 0.001) beta(2)-AR mRNA but had no effect (P > 0.10) on beta(1)-AR or beta(3)-AR mRNA. We conclude that days on feed and RAC are affecting beta-AR mRNA levels, which could, in turn, impact the biological response to RAC feeding in yearling steers.     

Alterations of Endothelium-Dependent Digital Vascular Responses in Horses Given Low-Dose Endotoxin

Low doses of endotoxin cause vasoconstriction and hypoperfusion of the digit, small intestine, and cecum in horses. To determine the potential cause of these vascular alterations, in vitro vascular responses of palmar digital arteries and veins were determined in 8 horses after intravenous (IV) infusion of 1 L 0.9% NaCl (control) and 0.1 μg/kg Escherichia coli 055:B5 endotoxin in 1 L of 0.9% NaCl (endotoxin-treated). Vessels were surgically removed under general anesthesia, cut into 4-mm vascular rings, suspended in tissue baths, and attached to force displacement transducers for measurement of vascular tension. Cumulative concentration response curves to acetylcholine, bradykinin, nitroprusside, norepinephrine, 5-hydroxytryptamine (serotonin), and endothelin were determined. Maximal relaxation or contraction and the concentrations needed to produce 50% maximal relaxation or contraction were determined. Palmar digital arteries from endotoxin-treated horses relaxed significantly less in response to acetylcholine and bradykinin (endothelium-dependent), but not to nitroprusside (endothelium-independent) when compared with arteries from control horses. Digital arteries from endotoxin-treated horses also contracted significantly more with norepinephrine but less with serotonin. Digital veins responded less than digital arteries. In another study, vascular reactivity experiments documented that acetylcholine and bradykinin were endothelium-dependent vasodilators (endothelium-denuded vessels relaxed less than control vessels) in palmar digital vessels. Additionally, maximal relaxations for both vasodilators were significantly inhibited by N-nitro-L-arginine methyl ester (L-NAME), a nitric oxide antagonist, suggesting that acetylcholine and bradykinin cause relaxation through the nitric oxide pathway. The data from these studies indicate that low dose endotoxin impairs endothelium-dependent relaxation and augments adrenergic contraction of palmar digital arteries in horses.     

Techniques for evaluation of right ventricular relaxation rate in horses and effects of inhalant anesthetics with and without intravenous administration of calcium gluconate.

OBJECTIVES: To determine the most repeatable method for evaluating right ventricular relaxation rate in horses and to determine and compare effects of isoflurane or halothane with and without the added influence of intravenously administered calcium gluconate on right ventricular relaxation rates in horses. ANIMALS: 6 Thoroughbred horses from 2 to 4 years old. PROCEDURE: 6 models (2 for monoexponential decay with zero asymptote, 3 for monoexponential decay with variable asymptote, and 1 for biexponential decay) for determining right ventricular relaxation rate were assessed in conscious and anesthetized horses. The 2 methods yielding the most repeatable results then were used to determine right ventricular relaxation rates in horses anesthetized with isoflurane or halothane before, during, and after i.v. administration of calcium gluconate. Right ventricular pressure was measured, using a catheter-tip high-fidelity pressure transducer, and results were digitized at 500 Hz from minimum rate of change in ventricular pressure. RESULTS: 2 models that used monoexponential decay with zero asymptote repeatedly produced an estimate for relaxation rate and were used to analyze effects of anesthesia and calcium gluconate administration on relaxation rate. Isoflurane and halothane each prolonged right ventricular relaxation rate, with greater prolongation evident in halothane-anesthetized horses. Calcium gluconate attenuated the anesthesia-induced prolongation in right ventricular relaxation rate, with greater response obtained in isoflurane-anesthetized horses. CONCLUSIONS AND CLINICAL RELEVANCE: Right ventricular relaxation rate in horses is assessed best by use of a monoexponential decay model with zero asymptote and nonlinear regression. Intravenous administration of calcium gluconate to isoflurane-anesthetized horses best preserves myocardial relaxant function.     

Are so many adrenergic receptor subtypes really present in domestic animal tissues? A pharmacological perspective

Adrenergic receptors (ARs) are the cellular membrane binding sites through which natural catecholamines and sympathomimetic drugs exert their physiological and pharmacological effects. In recent decades, studies to clarify the distribution and function of ARs have been performed mostly on cultured cells, laboratory animals and human target tissues, but little is known about these aspects in domestic animals. This review focuses on AR structure, classification and signalling pathways and on AR subtype distribution in target tissues of some domestic animals, namely dogs, horses and bovines. In these species, different alpha- and beta-AR subtypes have been characterized and the functions controlled by the adrenergic systems have been studied. In the dog, the role played by the adrenergic system in the pathogenesis of cardiovascular disorders and in the modulation of canine aggression has roused particular interest. In dogs affected by dilated cardiomyopathy a significant down-regulation of beta-ARs has been observed both in the heart and circulating lymphocytes. This finding confirms the involvement of the adrenergic system in the pathogenesis and progression of the disorder and suggests new therapeutic strategies. In the horse, AR distribution has been studied in the cardiac, respiratory and gastrointestinal systems as well as in digital veins and arteries. The cardiac beta-ARs in healthy horses seem to be predominantly represented by the beta(1) subtype. In this species, heart failure may increase the expression of the beta(2) subtype, rather than causing AR down-regulation. Different beta- and alpha-AR subtypes have been characterized in the smooth muscle of equine ileum. The sympathetic relaxation of equine ileum smooth muscle seems to depend mainly on beta(3)-AR subtype activation, with minor involvement of the beta(2) subtype. In the respiratory tract, regional differences have been evidenced in the functionality of beta-AR subtype. The beta(2) subtype predominates in all segments but the beta(2) subtype-mediated adenyl cyclase response is tissue-dependent, with higher activity in tracheal membranes than bronchial or pulmonary ones. Both alpha- and beta-AR subtypes are present in the genital tract of cows. Bovine ovarian and myometrial cell membranes express higher concentrations of beta(2)-ARs than the beta(1) subtype, whereas as far as alpha-ARs are concerned, a single class of alpha(1)-ARs and two distinct classes of alpha(2)-AR binding sites have been discriminated. Interestingly, it has been observed that the activation of the sympathetic system could play an important role in the pathogenesis of bovine ovarian cysts as suggested by the modifications in beta-AR levels in the hypophysis and ovary of cows affected by ovarian cysts. In this species, the phenomenon of down-regulation has been well studied in different organs of veal calves treated with clenbuterol as a "partitioning agent". Since differences exist in AR distribution among species, data obtained in laboratory animals or in human beings cannot be extrapolated to domestic animals and further investigation on AR subtypes in domestic animal tissues is necessary.     

Ontogenesis of mRNA levels and binding sites of hepatic alpha-adrenoceptors in young cattle

Catecholamines affect hepatic glucose production through (alpha- and beta2-) adrenoceptors (AR). We studied mRNA abundance and binding of hepatic alpha-AR in pre-term (P0) calves and in full-term calves at day 0 (F0), day 5 (F5) and day 159 (F159) to test the hypothesis that gene expression and numbers of hepatic alpha-AR in calves are influenced by age and associated with beta2-AR and selected traits of glucose metabolism. mRNA levels of alpha1- and alpha2-AR were measured by real time RT-PCR. alpha1- and alpha2-AR numbers (maximal binding, Bmax) were determined by saturation binding of (3H)-prazosin and (3H)-RX821002, respectively. alpha1- and alpha2-AR subtypes were evaluated by competitive binding. alpha1A-AR mRNA levels were lower in P0 than in F0, F5 and F159 and alpha(2AD)-AR mRNA levels were lower in F159 than in P0, F0 and F5, while alpha2C-AR mRNA levels increased from P0 and F0 to F5 and F159. Bmax of alpha1-AR increased from P0 to F5, then decreased in F159. Bmax of alpha2-AR decreased from F0 to F159. Bmax of alpha1-AR was positively associated with mRNA levels of alpha1A-AR (r = 0.7), Bmax of beta2-AR (r = 0.5) and negatively with hepatic glycogen content (r = -0.6). Bmax of alpha2-AR was negatively associated with Bmax of beta2-AR (r = -0.4). In conclusion, mRNA levels and binding sites of alpha1- and alpha2-AR in calves exhibited developmental changes and were negatively associated with hepatic glycogen content.