Direct effect of hypothalamic neuropeptides on the release of catecholamines by adrenal medulla in sheep - study ex vivo.

Stress causes the activation of both the hypothalamic-pituitary-adrenocortical axis and sympatho-adrenal system, thus leading to the release from the adrenal medulla of catecholamines: adrenaline and, to a lesser degree, noradrenaline. It has been established that in addition to catecholamines, the adrenomedullary cells produce a variety of neuropeptides, including corticoliberine (CRH), vasopressin (AVP), oxytocin (OXY) and proopiomelanocortine (POMC) - a precursor of the adrenocorticotropic hormone (ACTH). The aim of this study was to investigate adrenal medulla activity in vitro depending, on a dose of CRH, AVP and OXY on adrenaline and noradrenaline release. Pieces of sheep adrenal medulla tissue (about 50 mg) were put on 24-well plates and were incubated in 1 mL of Eagle medium without hormone (control) or supplemented only once with CRH, AVP and OXY in three doses (10-7, 10-8 and 10-9 M) in a volume of 10 µL. The results showed that CRH stimulates adrenaline and noradrenaline release from the adrenal medulla tissue. The stimulating influence of AVP on adrenaline release was visible after the application of the two lower doses of this neuropeptide; however, AVP reduced noradrenaline release from the adrenal medulla tissue. A strong, inhibitory OXY effect on catecholamine release was observed, regardless of the dose of this hormone. Our results indicate the important role of OXY in the inhibition of adrenal gland activity and thus a better adaptation to stress on the adrenal gland level.

Verapamil - L type voltage gated calcium channel inhibitor diminishes aggressive behavior in male Siamese fighting fish.

Verapamil is a L-type voltage gated calcium channels inhibitor (VGCCI), which is a highly prescribed drug used in the treatment of hypertension, angina pectoris, cardiac arrhythmia and cluster headaches. Its common use caused its appearance in water environment. VGCC inhibit epinephrine release and cause many neuro-hormonal changes influencing also fish behavior. Siamese fighting fish was chosen to study the influence of verapamil given to the water on the beginning of experiment in 3 different concentrations of 0 (control), 8 and 160 µg · L-1, on aggressive behavior in these fish. The experimental fish were placed in individual glass containers for 3 weeks and the mirror test was used. The highest concentration led to a significant modulation of fish behavior after 1 week and the lower dose caused statistically significant behavioral changes after 2 weeks of verapamil treatment. Siamese fighting fish males exposed to verapamil had longer latencies to the first chase - 12.6 s (8 µg · L-1 of verapamil) and 18.8 s (160 µg · L-1 of verapamil) compared to 5.6 s in the control group, decreased attack frequency and shorter duration of these attacks. The number of attacks within 10 min was decreased from 38.3 in the control group to 27.1 and 16.1, respectively. Also the total duration of these attacks decreased from 354.8 (control) to 326.4 (decrease statistically insignificant) and to 194.8 s in verapamil treated groups. It was shown, that even relatively low concentrations of verapamil in water may have adverse effects on fish and probably other living organisms.

Influence of centrally administered diltiazem on behavioural responses, clinical symptoms, reticulo-ruminal contractions and plasma catecholamine level after experimentally induced duodenal distension in sheep.

A different role of L-type antagonists for voltage-gated calcium channels (VGCC) has been previously identified in different types of experimental and clinical pain in man and animals. Present study examined the role of VGCC blocker - diltiazem administered icv (0.25, 0.5, 1.0 and/or 2.0mg in toto) on the development of pain related symptoms, clinical signs, plasma catecholamine level and the inhibition of reticulo-rumen motility caused by 5min lasting mechanical duodenum distension (DD) in the sheep. Experimental DD was conducted by insertion (during surgery) of rubber balloon into the duodenum and the distension by 40ml of warm water. Duodenal distension resulted in a significant increase of behavioural pain responses, tachycardia, hyperventilation, inhibition of reticulo-rumen contractions rate (from 85% to 45% during 15-20min), an increase of plasma catecholamine concentration (over sevenfold increase of epinephrine during 2h following DD, two-times norepinephrine and 84% increase of dopamine). Diltiazem infusion given 10min before DD decreased intensity of visceral nocifensive responses such as: behavioural changes, tachycardia, hyperventilation, reticulo-rumen motility and efficiently prevented appearance of catecholamine release. These data demonstrated that the development and persistence of acute duodenal pain depends on the activation of Ca(2+) ion flux leading to neurotransmitters release and modulation of membrane excitability. It seems that diltiazem given icv 10min prior to DD (as a source of acute visceral pain), inhibited specific receptors α(1) subunits of VGCCs in target tissues, prevent depolarization of cell membranes and release of neurotransmitters responsible for pain sensitivity in sheep. The observed antinociceptive action of VGCCs type-L blockers suggests that these channels play a crucial role in the modulation of acute visceral pain in sheep.

The inhibition of experimentally induced visceral hyperalgesia by nifedipine - a voltage-gated Ca(2+) channels blocker (VGCCs) in sheep.

Present study examined the effect of VGCC L-type blocker - nifedipine given i.c.v. (0.25, 0.5, 1 and/or 2mg in toto) on the development of nociceptive behavior, clinical symptoms, plasma catecholamin concentration and reticulo-rumen motility following 5 min lasting mechanical duodenal distension (DD) in sheep. After 24h of fasting, all animals received i.m. ketamine analgesia (20 mg kg(-1)B.W) and anesthetized with pentobarbital (20 mg kg(-1)B.W., i.v. infusion) The permanent stainless steel cannula 29 mm in length and 2mm in diameter was inserted into the lateral cerebral ventricle (controlled by cerebro-spinal efflux) 10mm above the bregma and 5mm laterally from the midline sutures using stereotaxic method. Under the same general anesthesia/analgesia a T-shaped silicon cannula (inside diameter of 21 mm), was inserted into the duodenum (12 cm from pylorus). Second identical cannule was inserted into the dorsal sac of the rumen, a previously described. After surgery each animal was kept in individual boxes for 10 days prior to experiment and was treated i.m. with benzyl procaine penicillin 30,000 I.U kg(-1)B.W.)+dihydrostreptomycine sulfate (10 g kg(-1)B.W.)+prednisolone acetate (1.2 mg kg(-1)B.W.) combination and i.m. ketamine (20 mg kg(-1)B.W.) every day by seven consecutive days. Experimental DD was conducted by insertion and then distension of rubber balloon (containing 40 ml of warm water) inserted into sheep duodenum. Duodenal distension produced a significant increase in behavioral pain manifestations, tachycardia, hyperventilation, inhibition of reticulo-ruminal contractions rate (from 87.2 to 38.0% during 15-20 min), an increase of plasma catecholamine concentration (over 6.4-fold increase of epinephrine during 2h following DD, 2-times norepinephrine and 84% increase of dopamine). Nifedipine infusion administered 10 min prior to DD decreased intensity of visceral pain manifestations such as: behavioral changes, hyperventilation, reticulo-rumen motility and efficiently prevent appearance of catecholamine release. These data demonstrated that the development and persistence of duodenal hyperalgesia depends on the activation of Ca(2+) ion flux leading to neurotransmitters release and modulation of membrane excitability. It seems that nifedipine given i.c.v. 10 min prior to DD (as a source of visceral pain), inhibited specific receptors 1 subunits of VGCCs in target tissues, prevented depolarization of cell membranes and release of neurotransmitters responsible for pain sensitivity in sheep. The observed antinociceptive action of VGCCs type L blockers suggest that these channels play a crucial role in the modulation of acute visceral hyperalgesia in sheep.

Centrally administered verapamil prevents the autonomic reaction to visceral pain in sheep.

The significant role of voltage gated calcium channels (VGCC) L-type antagonists used concomitantly with opioids in attenuation of clinical pain has been confirmed. The aim of this study was to evaluate the effect of centrally administered verapamil on behavior and biochemical parameters in sheep that have undergone experimental duodenal distension (DD) and to determine whether verapamil exerts any anti-nociceptive effects under these conditions. The study was carried out using 24 mature crossbred ewes, each weighing 38-43 kg. Verapamil, a VGCC blocker, was administered through an intracerebroventricular cannula at the following doses: 0.25, 0.5, 1.0 and 2.0mg in toto. Ten minutes later experimental DD was conducted by insertion and the distension of rubber balloon (containing 40 ml of warm water) inserted into sheep duodenum. After 5 min of mechanical DD the following reactions were then observed: the significant increase in behavioral pain responses, i.e. tachycardia, hyperventilation, inhibition of reticulo-ruminal contractions (70% approximately, during 15 min), an increase of plasma catecholamine concentration (over 7-fold increase of epinephrine during 2h following DD, 2-times norepinephrine and +/-80% increase of dopamine). Verapamil infusion administered 10 min prior to DD decreased intensity of visceral pain responses, such as: behavioral changes, tachycardia, hyperventilation, inhibition of the reticulo-rumen motility and efficiently prevented the appearance of catecholamine release. These data demonstrated that the development and persistence of duodenal hyperalgesia depends on the activation of Ca(2+) ion flux leading to neurotransmitters release and modulation of membrane excitability. The observed antinociceptive action of VGCCs type-L blockers suggests that these channels play a crucial role in the modulation of acute visceral hyperalgesia in sheep.

Influence of nifedypine on the hyperalgesic action of duodenal distention in sheep.

The aim of this study was to determine the influence of nifedypine--competitive antagonist of voltage-gated dependent L-type Ca2+ channels (VGCCs)--on inhibition of reticulo-ruminal motility, heart beats, respiratory rates and other nociceptive behavior symptoms caused by duodenal distention (DD). The animals, which were under general anesthesia, had duodenal and ruminal fistulas and intracerebroventriculary (i.c.v.) cannulas inserted into the lateral ventricle. Reticulo-ruminal contractions were recorded mechanographically using an electronic tensometer. The frequency of reticulo-ruminal contractions was determined by the number of mechanograms with 5 min intervals prior to and after DD (for 180 min). The duodenal distention was performed using a rubber balloon (10 cm length), which was inserted via the duodenal fistula and filled with 40 ml water. Five min DD caused immediate and almost complete inhibition of reticulo-ruminal contractions, nociceptive behavior symptoms, tachycardia and hyperventilation. Nifedypine per se did not change the reticulo-ruminal motility, general behavior or clinical symptoms; however, doses of 1 and 2 mg of nifedypine in toto infused i.c.v 10 minutes before DD prevented all signs of reticulo-ruminal disorders, as well as the general nociceptive behavior. Nifedypine inhibited particularly clinical symptoms such as tachycardia and hyperventilation. The observed antinociceptive action of VGCCs type-L blockers suggests that these channels play a crucial role in the modulation of acute visceral hyperalgesia. Nifedipine can be useful in controlling acute visceral pain associated, for example, with different kinds of colic.

Centrally administered PD 140.548 N-methyl-D-glucamine prevents the autonomic responses to duodenal pain in sheep.

Cholecystokinin (CCK) released in the CNS inhibits the analgesic action of exogenous opioids and may antagonize analgesia resulting from the activation of an endogenous pain inhibitory system. The aim of this study was to analyse the central action of PD 140.548 N-methyl-D-glucamine--a peptide antagonist of a specific peripheral type CCK receptor--on animal behaviour, catecholamines (CA) and cortisol concentration, as well as clinical symptoms of visceral pain induced by duodenal distension (DD). A 5 min distension of the duodenum wall, using a 10 cm long balloon filled with 40 and/or 80 ml of water (DD 40 and/or DD 80) at animal body temperature, produced a significant increase in plasma CA and cortisol levels, an increase in the heart rate, hyperventilation and other clinical symptoms (inhibition of rumen motility, bleating, teeth grinding, prostration, urination, defecation) that may be related to pain, proportionally to the degree of intestinal distension. Intracerebroventricular administration of PD 140.548 at the dose of 1 or/and 2 mg in toto 10 min before applying DD 40 completely blocked the increase in blood plasma cortisol, epinephrine (E), norepinephrine (NE) and dopamine (DA) concentration. It is suggested that the central inhibitory action of CCK antagonist on the cortisol and catecholamine release produced by visceral pain is due to the inhibition of peripheral CCK1 type receptors in the central centrifugal descending pain facilitatory system in sheep perhaps via the hypothalamic-pituitary-adrenal axis.

Intracerebroventricular administration of PD 140.548 N-methyl-D-glucamine attenuates the release of cortisol and catecholamines induced by duodenal distension in the sheep.

The aim of this study was to determine the influence of mechanically induced duodenal distension (DD) and PD 140.548 N-methyl-D-glucamine (a specific peptide antagonist of a CCK1 receptor) premedication on mechanographical reticulo-ruminal activity, animal general behaviour, catecholamines (CA) and the blood plasma cortisol levels, as well as the clinical symptoms of visceral pain induced by DD in sheep. After 24 h fasting, 6 animals, Polish merino sheep were praeanaesthetised by i.m. injection of ketamine (20 mg x kg(-1) b.w.) and anaesthetised with i.v. infusion of pentobarbital (20 mg x kg(-1) b.w.) and a permanent stainless steel cannula (gate cannula) was inserted inside the lateral cerebral ventricle (controlled by cerebrospinal fluid efflux) 10 mm above the bregma and 5 mm laterally from the midline suture using stereotaxic method. Under the same general anaesthesia and analgesia a T-shaped silicon cannula, was inserted into the duodenum (12 cm from pylorus) and a second one was inserted into the dorsal sac of the rumen. During 7 consecutive days after surgery each animal was treated i.m. with procaine penicillin (300000 I.U..kg(-1) b.w.), dihydrostreptomycine (DHS, 10 microg x kg(-1) b.w.), prednisolone acetate 1.2 mg x kg(-1) b.w.) together and i.m. injection of ketamine (20 mg x kg(-1) b.w.), separetely. The influence of PD 140.548 N-methyl-D-glucamine on the unfavourable effects of duodenal distension using a 10 cm long balloon filled with 40 and 80 ml (DD40 and DD80) water at animal body temperature was investigated in this study. Five minutes DD40 and DD80 caused an immediate and compete inhibition of the reticulo-ruminal frequency, a significant increase in plasma CA and cortisol levels, an increase in the heart rate, hyperventilation and other symptoms of pain, proportionally to the degree of intestinal distension. Intracerebroventricular (i.c.v.) administration of PD 140.548 alone at a dose of 0.25, 0.5, 1 or 2 mg in toto did not significantly change the reticulo-ruminal motility, CA and cortisol concentrations, but 10 min after the i.c.v. infusion (or 10 min before DD) at a dose 1 and 2 mg in toto , it completely blocked the increase of blood plasma cortisol, epinephrine (E), norepinephrine (NE) and dopamine (DA) concentrations for 20 min. In the some time it prevented the reticulo-ruminal atony provocked by DD. It is concluded that PD 140.548 N-methyl-D-glucamine--an antagonist of the central CCK1 receptor can be an effective analgesic agent in duodenal pain. This action is due to the inhibition of peripheral CCK1 type receptor in the central descending nerve pathway, facilitating pain transmission in sheep perhaps in the hypothalamic-pituitary-adrenal axis.

Pharmacological and toxicological aspects of combination of beta-lactam and aminoglycoside antibiotic, prednisolone and procaine hydrochloride on the example of Vetramycin.

Vetramycin is an injectable veterinary compound for animal use only. In veterinary medicine, it has been used for a long time as a bactericidal beta-lactam and aminglycoside antibiotics combination, extending the bactericidal spectrum of these substances. This compound, in addition to bactericidal procaine penicillin and dihydrostreptomycin (DHS), contains also prednisolone acetate and procaine hydrochloride, two biologically active substances. Prednisolone, a glucocorticoide, has an antiinflammatory, antiallergic, antiitchical and analgesic effect. Procaine hydrochloride, in turn, has a local anaesthetic effect and attenuates pain caused by irritable properties of antibiotics at the injection sites. The average dosage of, respectively, procaine benzylpenicillin (I.U./kg(-1) b.w.), DHS (microg/kg(-1) b.w.), prednisolone acetate (microg/kg(-1) b.w.) and procaine hydrochloride (mg/kg(-1) b.w.) in horses, cattle, pigs is 6000-15000, 10-11, 0.24-0.6 and 1.2-3.0; s.i.d., in sheep, foals, calves, piglets is 20000-40000, 10, 0.8-1.6 and 4-8; s.i.d., in dogs and cats is 30000-200000, 10, 0.8-1.6 and 4-8; s.i.d.. Intramammary injection dose (Vetramycin antimastitis ointment in syringe) in cows is 1000000 I.U. of procaine benzylpenicillin + 1000000 I.U. of dihydrostreptycin sulphate per quarter of udder, s.i.d., during 3 successive days.

CCK1 central receptor antagonist prevented the intestinal stress symptoms in sheep.

The aim of this study was to determine the influence of mechanically induced duodenal distension (DD) and lorglumide (CCK1 receptor antagonist) premedication on electrical activity of various parts of gastrointestinal (GI) tract and the blood plasma cortisol level in sheep. The influence of lorglumide on the unfavourable effects of duodenal distension (performed with a balloon filled with water--40 and 80 ml; DD40 and DD80) was investigated in this study. These effects in sheep were as follows: the atony of forestomachs and abomasum and the transitory stimulation of myoelectrical activity of small intestine and distal parts of large intestine. The animals, under general anaesthesia, had electrodes inserted into the muscular layers of the organ, the duodenal fistula and (in another group of animals) also the ruminal fistula. Five minute duodenal distension (DD40 and DD80) caused an immediate and complete inhibition of the frequency of spike bursts as well as reticulo-ruminal and abomasal contractions, but also a transitory significant increase of spike bursts of the intestinal wall. The duodenal distension (DD40 and DD80) caused a significant increase of plasma cortisol concentration. Lorglumide did not significantly change the motility of gastrointestinal tract and cortisol concentration, but 10 min after the intracerebroventricular (i.c.v.) infusion in the doses of 1 and 2 mg in toto (i.e. 25 and 50 micrograms/kg B.W.) it decreased the cortisol concentration by 59.7%, as compared with the control values. Lorglumide administered in the above mentioned doses 10 min before the DD40 prevented all signs of intestinal stress and decreased the release of cortisol, but only for 10 min since the beginning of the duodenal distension. It is concluded, that lorglumide--an antagonist of the central CCK1 receptors can be an effective antistressoric agent in the stomach atony caused by the duodenal distension (mechanical-algetic-emotional stress) in sheep.

The evaluation of the role of CCK in the opioid modulation of the motility of the gastrointestinal tract in sheep.

The participation of central cholecystokinin-8 (CCK-8) receptors in the modulatory effect of D-Ala2, N-Me-Phe4, Gly5-ol enkephalin (DAGO), a selective mu-opioid receptor agonist, on the spike burst activity of the gastrointestinal tract (rumen, reticulum, antrum, duodenum, colon and caecum) in sheep was investigated. DAGO was infused intracerebroventricularly (i.c.v.) at doses of 0.1-1 microg/kg body weight (BW). It was shown that DAGO significantly inhibited myoelectrical activity of the wall of the forestomachs, abomasum and colon but stimulated this activity in the duodenum (rate of myoelectrical migrant complex-MMC). The effects of DAGO were prevented by CCK-8 antagonists (L-364.718 and L-365.260) previously infused at doses of 5-20 microg/kg BW. The results of this present study indicate that central receptors of CCK-8 participated in the modulatory action of an opioid on myoelectrical activity of the gastrointestinal tract in sheep. Furthermore, this result suggests that CCK-8 is released in response to mu-receptor stimulation, because CCK-8 antagonists (L-364.718 and L-365.260) prevented the modulatory action of DAGO on the gastrointestinal motility in sheep.

Hypothalamus involvement in the reticulo-rumen motor and behavioural disturbances induced by morphine in sheep.

Morphine (20 and 40 micrograms/kg) administered into the cerebral ventricle of conscious sheep caused significant inhibition of the mean frequency and the average amplitude of primary ruminal contractions by 45 min after injection. Between 90 and 120 min, morphine (40 micrograms) provoked a significant increase in the amplitude (p < 0.01). At both doses it caused strong psychomotor excitability that lasted for more than 140 min. Isolation of the hypothalamus prevented both the inhibitory effects of morphine on rumen motility and the drug-induced psychomotor excitability. Histopathological analysis of slices of the hypothalamus, pons and medulla indicated descending degenerative changes in the nervous pathways connecting the hypothalamus with lower structures in the brain. These results suggest either that hypothalamic isolation caused degeneration of inhibitory descending pathways that connect the hypothalamus with the gastric centres or that structures of importance for forestomach motility are not located within the gastric centres but elsewhere in the brain, for example in the hypothalamus.

Central and peripheral serotonergic influences on viscerovisceral inhibitory reflex during duodenal distension in sheep.

The effects of duodenal distension on forestomach and abomasal motility were investigated in conscious sheep chronically fitted with intraparietal electrodes, a duodenal cannula, and an intracerebroventricular cannula. Duodenal distensions with a balloon inflated with 40 ml (DD40) of water reduced the frequency of forestomach and abomasal contractions by 45 and 32%, respectively, while distension with 80 ml (DD80) induced a total inhibition. Methysergide, a mixed 5HT1-5HT2 antagonist administered intravenously (200 micrograms/kg) or intracerebroventricularly (20 micrograms/kg) suppressed the DD40-induced inhibition and reduced that induced by DD80. Spiroxatrine, a selective 5HT1A antagonist, intravenously (100 micrograms/kg) or intracerebroventricularly (10 micrograms/kg), suppressed the DD40 and DD80-induced inhibition, which was also attenuated by the 5HT2 antagonist ritanserin given intravenously (200 micrograms/kg) or intracerebroventricularly (20 micrograms/kg). Granisetron, a 5HT3 antagonist, injected intravenously (150 micrograms/kg), abolished the effects of DD40 and DD80 while it had no antagonistic action on DD40 and DD80 when given intracerebroventricularly (15 micrograms/kg). It is concluded that in sheep, duodenal distension inhibits forestomach and abomasal motility through 5HT1A and 5HT2 receptors at the level of the central nervous system and 5HT3 receptors located peripherally.

Beta-endorphin-induced inhibition of rumen contractions in sheep. The effect of hypothalamic de-efferentiation.

In conscious sheep, beta-endorphin (1 and 2 micrograms/kg) administered into the third cerebral ventricle caused psychomotor excitability and a significant inhibition of the frequency of rumen contractions. The amplitude of the first rumen contractions, following immediately after the end of endorphin infusion, and the average amplitude of primary rumen contractions were also inhibited. De-efferentiation at the level of the hypothalamus prevented both the inhibitory effect of beta-endorphin on the frequency of rumen contractions and the drug-induced psychomotor excitability. However, de-efferentiation did not prevent beta-endorphin-induced inhibition of the mean amplitude of rumen contractions. The character of pathohistological changes induced by de-efferentiation showed descending degenerative changes of the nerve tracts connecting the hypothalamus with the pons and the medulla oblongata. These results, together with previously published evidence, do suggest that de-efferentiation at the level of the hypothalamus causes degeneration of inhibitory descending opioid-noradrenergic pathways connecting the hypothalamus with the gastric centres in the medulla oblongata.

Opioid inhibitory control of the ruminant stomach motility: functional importance of the hypothalamus.

In conscious sheep, Leu-enkephalin (10 and 20 micrograms.kg-1) administered into the third cerebral ventricle caused a psychosedative action and a significant inhibition of the frequency of rumen contractions. The amplitude of the first rumen contractions, following immediately after the end of the Leu-enkephalin's infusion (at a dose of 20 micrograms) and the average amplitude of primary rumen contractions were also significantly inhibited. The hypothalamic isolation prevented both the inhibitory effect of Leu-enkephalin on the rumen motility and the drug induced psychosedative action. In contrast, Leu-enkephalin (20 micrograms.kg-1), significantly enhanced phasic contractions of the rumen after hypothalamic isolation. However the hypothalamic isolation did not prevent Leu-enkephalin induced inhibition of the mean amplitude of rumen contractions, but this inhibition was not significant (NS). The character of the histopathological changes induced by the isolation of the hypothalamus showed descending degenerative changes of the neuronal pathways connecting the hypothalamus to the pons and lower to the medulla. These results do suggest that the isolation of the hypothalamus causes the degeneration of the descending inhibitory opioid pathways connecting the hypothalamus to the gastric centers of the medulla and point act out the functional importance of descending tonic opioid inhibitory control of ruminant forestomach motility.

Modification by 6-hydroxydopamine (6-OHDA) of the inhibition of extrinsic rumen contractions in sheep produced by beta-endorphin.

In sheep, beta-endorphin (1 and 2 micrograms/kg) administered into the third cerebral ventricle caused a significant inhibition of the frequency of rumen contractions. The amplitude of the first rumen contractions, following immediately after the end of infusion, and the average amplitude of primary rumen contractions, were inhibited. Beta-endorphin caused general psychomotor excitability. These results suggest that an inhibitory mu and delta opioid system is involved in the control of forestomach motility and general behaviour in sheep. All effects of beta-endorphin were completely prevented by i.c.v. 6-hydroxydopamine (6-OHDA, 18.2 micrograms/kg) pre-treatment. These results suggest that beta-endorphin-induced inhibition of rumen motility is due to central noradrenergic system activation. The exact location of this noradrenergic system remains to be determined.

The effect of central sympathectomy by 6-hydroxydopamine (6-OHDA) on the response to morphine in conscious sheep.

When morphine, an opioid mu-agonist, was administered in vivo into the third cerebral ventricle (ICV) of conscious sheep at 20 and 40 micrograms/kg body weight, it caused psychomotor excitability for 2-3 h and a significant decrease in the reticuloruminal frequency for 45 min and in the mean amplitude of the primary contractions for 65 min. From 60 min after infusion, the same doses of morphine caused a significant increase in the average amplitude of the contractions for 45 min. This suggests that an inhibitory mu-opioid acceptor is involved in the central control of forestomach motility and general behaviour in sheep. All the effects of morphine were completely prevented by pretreatment with 18.2 micrograms/kg body weight 6-OHDA ICV. These results suggest that both morphine-induced inhibition of rumen motility and psychomotor excitability are due to central noradrenergic descending system activation. The exact location of the noradrenergic system remains to be determined.

Neurochemical changes in the brain and spinal cord of sheep: a basis for the immobilizing action of etorphine.

Clinical and pharmacological analyses showed a full immobilizing action of etorphine in adult sheep +/- 7 min. after a deep i.m. injection of the drug at a dosage rate of 20 micrograms/kg. Neurochemical analyses of motoric CNS-structures done within 30 min. after i.m. injection of etorphine in an immobilizing dose, showed the following: A significant decrease of dopamine (DA) and homovanilic acid (HVA) concentrations in the corpus striatum, the frontal motor cortex, cerebellum and in the lumbo-sacral portion of the spinal cord. A decrease of DA-concentration with a simultaneous increase of HVA-concentration in the pons. A significant decrease of noradrenaline (NA) concentration in the cerebellum and lumbo-sacral portion of the spinal cord and an insignificant decrease of amine concentration in the pons. A significant increase of adrenaline (A) concentration in the frontal motor cortex. A significant decrease of 5-hydroxytryptamine (5-HT) concentration in the frontal motor cortex and cerebellum and an equally significant increase in its concentration in the pons.

Presynaptic stimulation of dopaminergic CNS structures in sheep as a mechanism of immobilising action of Immobyl (fentanyl + azaperone).

The immobilising action of Immobyl (fentanyl : azaperone, 5:1) in six sheep has been analysed on the basis of the changes in dopamine, noradrenalin, adrenalin, homovanillic acid and 5-hydroxytryptamine concentrations in the corpus striatum, frontal motor cortex, pons, cerebellum and lumbosacral spinal cord as compared to the control animals which were given saline. Forty minutes after intramuscular injection of an immobilising dose (0.19 mg [kg bodyweight]-1), Immobyl caused a significant decrease in dopamine, noradrenalin and 5-hydroxytryptamine concentrations and a similarly large decrease in homovanillic acid concentration (47 per cent) in the corpus striatum with a simultaneous but insignificant increase in the concentrations of these substances in the frontal motor cortex region. In Immobyl immobilisation, sheep showed a significant increase in dopamine concentration with an equally significant decrease in homovanillic acid concentrations in the lumbosacral part of the spinal cord. It is suggested that fentanyl stimulates the presynaptic dopamine receptors in the corpus striatum in sheep, significantly decreasing synthesis and release of dopamine and noradrenalin and intensifying an inhibitory effect of the corpus striatum on locomotor activity and thus causes the immobilisation of the animal.

Inhibitory action of intravenously administered ammonium acetate on the motility of the rumen in sheep.

The effect of 60 minutes' intravenous infusions, before morning feeding, of ammonium acetate (18.6 micromole/min/kg of body weight) and ammonium acetate with propranolol (11 microgram/min/kg of body weight) on the ruminal motility of sheep was examined. Ammonia has a adrenaline-like action therefore propranolol, a beta-receptor blocking agent, was administered in order to eliminate the possible effect of adrenaline on ruminal motility. The contractions of the dorsal sack of the rumen were registered by means of the balloon method, with the gauge inserted through the rumen fistula. The infusion of ammonium acetate caused an increase of the ammonia concentration in the blood to 0.6 mmol/1 at the end of 60 minutes' infusion. Already during the first 5 minutes of the intravenous infusion of ammonium acetate there was a decreased frequency of ruminal contractions, which was observed throughout the infusion. After the infusion there was a radical decrease of the concentration of ammonia in the blood, and at the same time an increase in the frequency of rumen contraction was observed. Blocking of the beta-adrenergic receptors by propranolol did not eliminate the inhibiting action of ammonium ion on rumen motility. The infusion of the ammonium acetate caused an increase of adrenaline and glucose concentration. This response was eliminated by propranolol in the case of adrenaline but not glucose. It is assumed that the action of ammonium ion on the rumen motility is derived primarily by the central nervous system.