ABSTRACT
BACKGROUND: Discovery of adequate pharmacological treatments for constipation has proven elusive. Increased numbers of bowel movements were reported as a side-effect of ulimorelin treatment of gastroparesis, but there has been no investigation of the site of action. METHODS: Anesthetized rats were used to investigate sites and mechanisms of action of ulimorelin. KEY RESULTS: Intravenous ulimorelin (1-5 mg/kg) caused a substantial and prolonged (~1 h) increase in colorectal propulsive activity and expulsion of colonic contents. This was prevented by cutting the nerves emerging from the lumbosacral cord, by the nicotinic receptor antagonist hexamethonium and by antagonists of the ghrelin receptor. The effect of intravenous ulimorelin was mimicked by direct application of ulimorelin (5 µg) to the lumbosacral spinal cord. CONCLUSIONS & INFERENCES: Ulimorelin is a potent prokinetic that causes propulsive contractions of the colorectum by activating ghrelin receptors of the lumbosacral defecation centers. Its effects are long-lasting, in contrast with other colokinetics that target ghrelin receptors.
Subject(s)
Defecation/drug effects , Macrocyclic Compounds/pharmacology , Receptors, Ghrelin/agonists , Animals , Colon/drug effects , Colon/physiology , Injections, Spinal , Macrocyclic Compounds/administration & dosage , Male , Rats , Rats, Sprague-Dawley , Rectum/drug effects , Rectum/physiology , Spinal Cord/drug effects , Spinal Cord/physiologyABSTRACT
STUDY DESIGN: Animal proof of principle study. OBJECTIVES: To determine whether capromorelin, a compound that causes defecation by stimulating ghrelin receptors within the lumbosacral defecation centers, is effective after spinal cord injury (SCI), and whether SCI significantly alters sensitivity to the compound. SETTING: University of Melbourne and Austin Hospital, Melbourne, Australia. METHODS: Rats were subjected to spinal cord contusion injury or were sham-operated. At 6 weeks after surgery, effects of capromorelin on blood pressure, heart rate and propulsive contractions of the colorectum were investigated. RESULTS: Capromorelin caused robust propulsive activity in the colorectum soon after its application. The compound was similarly effective in naïve, sham-operated and spinal cord-injured rats. Blood pressure increases caused by capromorelin were not exaggerated after SCI, and there was no evidence of phasic blood pressure increases when the colon was contracted by the compound. CONCLUSION: Capromorelin is a therapeutic compound that could potentially be used to relieve constipation by triggering defecation in spinal cord-injured patients.
Subject(s)
Constipation/drug therapy , Constipation/physiopathology , Defecation/drug effects , Piperidines/pharmacology , Pyrazoles/pharmacology , Receptors, Ghrelin/agonists , Spinal Cord Injuries/physiopathology , Animals , Constipation/etiology , Defecation/physiology , Disease Models, Animal , Growth Hormone/metabolism , Male , Rats , Rats, Sprague-Dawley , Receptors, Ghrelin/physiology , Spinal Cord Injuries/complicationsABSTRACT
The pelvic autonomic nervous system is a target for circulating androgens in adults, with androgen exposure or deprivation affecting the structure and function of urogenital tract innervation. However, the critical period for androgen exposure to initially establish pelvic autonomic neuromuscular transmission has not been determined. We have examined the sympathetic innervation of the vas deferens in hypogonadal (hpg) mice that are deprived of androgens after birth but undergo normal prenatal sexual differentiation and remain androgen responsive throughout life. In vasa deferentia from hpg mice, purinergic excitatory junction potentials and contractions could not be elicited by electrical stimulation and P2X(1) purinoceptors could not be demonstrated by immunofluorescence. Moreover, a novel inhibitory nitrergic transmission developed. Administering testosterone to adult hpg mice restored purinergic excitatory transmission and P2X(1) purinoceptor immunofluorescence, and nitrergic inhibitory transmission was lost. Despite the deficit in excitatory neurotransmission in hpg mice, their vasa deferentia were innervated by numerous noradrenergic axons and pelvic ganglia appeared normal. In addition, noradrenergic contractions could be elicited by electrical stimulation. This study has revealed that postnatal androgen exposure has a profound effect on the development of excitatory transmission in vas deferens smooth muscle, primarily by a postjunctional action, but is not essential for development of the structural innervation of this organ. Our results also indicate that there is no postnatal critical period for androgen exposure to establish neuroeffector transmission and that postnatal androgen exposure can be delayed until adulthood, with little consequence for establishment of normal sympathetic neurotransmission.
Subject(s)
Androgens/metabolism , Hypogonadism/physiopathology , Muscle Contraction , Muscle, Smooth/innervation , Sympathetic Nervous System/physiopathology , Synaptic Transmission , Vas Deferens/innervation , Acetylcholine/metabolism , Adenosine Triphosphate/analogs & derivatives , Adenosine Triphosphate/pharmacology , Age Factors , Aging/metabolism , Androgens/deficiency , Androgens/pharmacology , Animals , Disease Models, Animal , Electric Stimulation , Excitatory Postsynaptic Potentials , Hypogonadism/genetics , Hypogonadism/metabolism , Inhibitory Postsynaptic Potentials , Male , Mice , Mice, Mutant Strains , Muscle Contraction/drug effects , Muscle, Smooth/drug effects , Neural Inhibition , Nitrergic Neurons/metabolism , Norepinephrine/metabolism , Receptors, Purinergic P2/metabolism , Receptors, Purinergic P2X , Sympathetic Nervous System/drug effects , Sympathetic Nervous System/metabolism , Synaptic Transmission/drug effects , Testosterone/metabolism , Time Factors , Vas Deferens/drug effectsABSTRACT
1. Intracellular recording was used to investigate the electrophysiological effects of activating peptidergic primary afferent axons with capsaicin in the smooth muscle of rat mesenteric arteries in vitro. In addition, continuous amperometry was used to monitor the effects of capsaicin on noradrenaline release from the sympathetic nerves. 2. Capsaicin (1 microm) produced a hyperpolarization (-11+/-2 mV) and a reduction in the time constant of decay of excitatory junction potentials (e.j.p.'s) evoked by electrical stimulation of the perivascular sympathetic nerves. These effects of capsaicin were mimicked by calcitonin gene-related peptide (CGRP; 1 and 10 nm) but not by substance P (50 nm), which produced a small hyperpolarization (maximum -3+/-1 mV) but did not change excitatory junction potential (e.j.p.) time course. 3. The hyperpolarization produced by capsaicin and CGRP was blocked by glibenclamide (10 microm) but was not changed by the CGRP antagonist, CGRP8-37 (0.5 microm). Mechanical denudation of the endothelium also did not reduce the effect of capsaicin on membrane potential. 4. Capsaicin (1 microm) increased the amplitude of e.j.p.'s. This effect was not mimicked by CGRP or substance P nor blocked by glibenclamide or CGRP8-37. 5. All effects of capsaicin desensitized. 6. Capsaicin (1 microm) had no effect on noradrenaline-induced oxidation currents evoked by electrical stimulation, indicating that noradrenaline release was unchanged. 7. These results suggest that CGRP released from primary afferent axons hyperpolarizes vascular smooth muscle by activating glibenclamide-sensitive K+ channels. The findings also indicate that an unknown factor released by the primary afferent axons increases e.j.p. amplitude.
Subject(s)
Capsaicin/pharmacology , Mesenteric Arteries/drug effects , Neurons, Afferent/drug effects , Animals , Apamin/pharmacology , Calcitonin Gene-Related Peptide/metabolism , Calcitonin Gene-Related Peptide/pharmacology , Electric Stimulation , Electrophysiology , Endothelium, Vascular/physiology , Female , Glyburide/pharmacology , In Vitro Techniques , Membrane Potentials/drug effects , Mesenteric Arteries/innervation , Mesenteric Arteries/physiology , Neurons, Afferent/metabolism , Neurons, Afferent/physiology , Norepinephrine/pharmacology , Peptide Fragments/pharmacology , Potassium Channel Blockers/pharmacology , Rats , Rats, Wistar , Substance P/pharmacology , Vasoconstriction/drug effects , Vasoconstrictor Agents/pharmacologyABSTRACT
1. Extracellular recording techniques were used to study nerve terminal impulses (NTIs) recorded from single polymodal nociceptors and cold-sensitive receptors in guinea-pig cornea isolated in vitro. 2. The amplitude and time course of NTIs recorded from polymodal nociceptors was different from those of cold-sensitive receptors. 3. Bath application of tetrodotoxin (1 microM) changed the time course of spontaneous NTIs recorded from both polymodal and cold-sensitive receptors. 4. Bath application of lignocaine (lidocaine; 1-5 mM) abolished all electrical activity. 5. Local application of lignocaine (2.5 and 20 mM) through the recording electrode changed the time course of the NTIs recorded from polymodal nociceptors but not that of NTIs recorded from cold-sensitive nerve endings. 6. It is concluded that action potentials propagate actively in the sensory nerve endings of polymodal nociceptors. In contrast, cold-sensitive receptor nerve endings appear to be passively invaded from a point more proximal in the axon where the action potential can fail or be initiated.
Subject(s)
Cold Temperature , Cornea/innervation , Nociceptors/physiology , Presynaptic Terminals/physiology , Thermoreceptors/physiology , Action Potentials/drug effects , Action Potentials/physiology , Anesthetics, Local/pharmacology , Animals , Evoked Potentials/drug effects , Evoked Potentials/physiology , Guinea Pigs , Lidocaine/pharmacology , Tetrodotoxin/pharmacologyABSTRACT
This study investigated the effects of changing the extracellular Ca2+ concentration on facilitation of excitatory junction potential (EJP) amplitude during trains of 20 stimuli at 1 Hz at sympathetic neuroeffector junctions in the guinea-pig vas deferens. These effects were compared with those of Ca2+ channel blockers and agents which act at prejunctional receptors to increase or decrease neurotransmitter release. In these experiments, alpha-adrenoceptor-mediated autoinhibition of neurotransmitter release was blocked by the alpha-adrenoceptor antagonist, phentolamine (1 microM). Varying the extracellular Ca2+ concentration (0.75-6 mM) changed the amplitude of EJPs without affecting the maximum level of facilitation during the trains of stimuli. Reductions in Ca2+ concentration (from 2 mM) were associated with a slowing in the rate of development of facilitation. The Ca2+ channel antagonists, Cd2+ (2 microM and 5 microM) and omega-conotoxin GVIA (10 nM), and agents which act at prejunctional receptors to reduce neurotransmitter release, adenosine (100 microM and 1,000 microM) and prostaglandin E2 (PGE2; 0.1 nM and 1 nM), produced similar effects to those of lowering the extracellular Ca2+ concentration. Raising the extracellular Ca2+ concentration (from 2 mM) increased the rate of development of facilitation. Angiotensin II (AII; 0.5 microM) produced similar effects to raising extracellular Ca2+. However, isoprenaline (1 microM), while increasing EJP amplitude, reduced the maximum level of facilitation and was without effect on the rate of development of facilitation. In the guinea-pig vas deferens EJPs are produced by neurally released ATP. Thus, the findings support the idea that adenosine, PGE2 and AII change ATP release by modifying Ca2+ entry into the nerve terminal. However, the effects of isoprenaline may not solely be accounted for by modifications in Ca2+ entry.
Subject(s)
Calcium/metabolism , Receptors, Presynaptic/metabolism , Synaptic Transmission/physiology , Vas Deferens/physiology , Adrenergic beta-Agonists/pharmacology , Angiotensin II/pharmacology , Animals , Calcium Channel Blockers/pharmacology , Dose-Response Relationship, Drug , Guinea Pigs , Isoproterenol/pharmacology , Male , Presynaptic Terminals/drug effects , Presynaptic Terminals/physiology , Synaptic Transmission/drug effects , Vas Deferens/drug effects , Vasoconstrictor Agents/pharmacologyABSTRACT
1. This study investigated the effects of BIIE0246, a novel neuropeptide Y (NPY) Y2 receptor antagonist, on the inhibition of cholinergic neuroeffector transmission in rat heart and guinea-pig trachea and purinergic neuroeffector transmission in guinea-pig vas deferens produced by the NPY Y2 receptor agonist, N-acetyl [Leu28,31] NPY 24-36. 2. In pentobarbitone anaesthetized rats, supramaximal stimulation every 30 s, of the vagus nerve innervating the heart, increased pulse interval by approximately 100 ms. This response was attenuated by intravenous administration of N-acetyl [Leu28,31] NPY 24-36 (10 nmol x kg(-1)). 3. Transmural stimulation of segments of guinea-pig trachea at 1 min intervals with 5 s trains of stimuli at 0.5, 5, 10, 20 and 40 Hz evoked contractions which were reduced in force by N-acetyl [Leu28,31] NPY 24-36 (2 microM). 4. In guinea-pig vasa deferentia, the amplitude of excitatory junction potentials evoked by trains of 20 stimuli at 1 Hz was reduced in the presence of N-acetyl [Leu28,31] NPY 24-36 (1 microM). 5. In all preparations BIIE0246 attenuated the inhibitory effect of N-acetyl [Leu28,31] NPY 24-36 but had no effect when applied alone. 6. The findings support the view that the nerve terminals of postganglionic parasympathetic and sympathetic neurones possess neuropeptide Y Y2 receptors which, when activated, reduce neurotransmitter release.
Subject(s)
Arginine/analogs & derivatives , Arginine/pharmacology , Benzazepines/pharmacology , Neuroeffector Junction/drug effects , Receptors, Neuropeptide Y/antagonists & inhibitors , Acetylcholine/metabolism , Action Potentials/drug effects , Animals , Female , Guinea Pigs , Heart Rate/drug effects , In Vitro Techniques , Male , Neuroeffector Junction/physiology , Purines/metabolism , Rats , Rats, Wistar , Trachea/drug effects , Trachea/physiology , Vas Deferens/drug effectsABSTRACT
The effects of the cholinesterase inhibitor neostigmine on the responses to vagus nerve stimulation of isolated sinus venosus/atrial preparations of the toad, Bufo marinus, were examined. In control solutions, trains of stimuli applied to the vagus nerve led to a decrease in heart rate that was susceptible to muscarinic receptor blockade. Membrane potential recordings made from sinus venosus cells showed that the responses to trains of stimuli, delivered at frequencies of less than 10 Hz, were little changed by the addition of neostigmine. However, the responses to longer trains of stimuli at 10 Hz (30 versus 10 s) were potentiated and the nature of the membrane potential changes was altered. The results suggest that, due to the activity of cholinesterases, acetylcholine (ACh) released from parasympathetic nerves normally has little access to the muscarinic receptors in the pacemaker region, which are linked to potassium channels.
Subject(s)
Cholinesterase Inhibitors/pharmacology , Heart/innervation , Neostigmine/pharmacology , Vagus Nerve/drug effects , Vagus Nerve/physiology , Acetylcholine/physiology , Action Potentials/drug effects , Action Potentials/physiology , Animals , Barium/pharmacology , Bufo marinus , Electric Stimulation , Heart Atria/innervation , Hydrolysis , Parasympathetic Nervous System/drug effects , Parasympathetic Nervous System/physiology , Potassium Channels/metabolism , Receptors, Muscarinic/metabolismABSTRACT
This study used intracellular recording of excitatory junction potentials (EJPs) and focal extracellular recording of excitatory junction currents (EJCs) to investigate the effects of agents that modulate intracellular cAMP levels on sympathetic neuroeffector transmission in the guinea-pig vas deferens. In this tissue, postjunctional electrical activity is produced by neurally released ATP. The adenylate cyclase activator, forskolin (0.5-5 microM) increased the amplitude of all EJPs evoked by trains of 20 stimuli at I Hz. Forskolin (5 microM) also increased the probability of recording EJCs without changing the amplitude distributions of spontaneous EJP and EJCs, indicating that this agent does not change the postjunctional sensitivity to spontaneously released quanta of ATP. EJP amplitudes were also increased by 8-bromo-cyclic AMP (10 microM), 8-bromo-cyclic GMP (10 microM), the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (100 and 1,000 microM) and the beta-adrenoceptor agonist, isoprenaline (1 microM). The selective protein kinase A inhibitors, H-89 (10 microM) and the Rp isomer of adenosine-3',5'-cyclic monophosphorothioate (Rp-cAMPS, 100 microM), and the broad spectrum protein kinase inhibitors, [1-(5-isoquinolinesulphonyl)-3-methylpiperazine-diHCl (H-7, 100 microM) and staurosporine (1 microM), did not block the facilitatory effects of forskolin on EJP amplitude. In addition, the effects of forskolin were not blocked by the cyclic nucleotide-gated ion channel blocker, spermine (50 microM). These results suggest that elevating intracellular cAMP levels increases ATP release in the guinea-pig vas deferens by a mechanism which does not involve activation of protein kinases A or G.
Subject(s)
Cyclic AMP/metabolism , Neuromuscular Junction/drug effects , Vas Deferens/drug effects , Adenosine Triphosphate/metabolism , Animals , Colforsin/pharmacology , Cyclic AMP-Dependent Protein Kinases/antagonists & inhibitors , Cyclic AMP-Dependent Protein Kinases/metabolism , Drug Interactions , Enzyme Activation , Enzyme Inhibitors/pharmacology , Guinea Pigs , In Vitro Techniques , Male , Membrane Potentials/drug effects , Membrane Potentials/physiology , Neuromuscular Junction/metabolism , Neuromuscular Junction/physiology , Neurotransmitter Agents/metabolism , Neurotransmitter Agents/physiology , Spermine/pharmacology , Vas Deferens/metabolism , Vas Deferens/physiologyABSTRACT
Continuous amperometry was used to monitor noradrenaline (NA) release from sympathetic nerves supplying rat mesenteric arteries in vitro. During electrical stimulation the amplitude of oxidation currents evoked by successive stimuli varied over a small range, with occasional events of larger amplitude. In the absence of stimulation, spontaneous oxidation currents (s.o.cs) were recorded. The frequency of s.o.cs was increased by alpha-latrotoxin (1 nM). This agent also increased the frequency of spontaneous excitatory junction potentials (s.e.j.ps), which monitor the packeted release of adenosine 5' triphosphate (ATP). The frequency of s.o.cs recorded 20-25 min after applying alpha-latrotoxin was about four times the control value, but that of s.e.j.ps was about 30 times the control value. The findings suggest that continuous amperometry can detect the spontaneous packeted release of NA, probably from large dense-cored vesicles. In contrast, s.e.j.ps monitor spontaneous release of neurotransmitter (ATP) from a different store, most likely the small dense-cored vesicles.
Subject(s)
Mesenteric Arteries/physiology , Nerve Endings/metabolism , Norepinephrine/metabolism , Sympathetic Nervous System/metabolism , Animals , Electric Stimulation , Electrochemistry , Electrophysiology , Evoked Potentials/drug effects , Female , In Vitro Techniques , Mesenteric Arteries/innervation , Mesenteric Arteries/metabolism , Muscle, Smooth, Vascular/innervation , Muscle, Smooth, Vascular/physiology , Oxidation-Reduction , Rats , Rats, Wistar , Spider Venoms/pharmacology , Time FactorsABSTRACT
1. Characteristic features of noradrenaline (NA) and adenosine 5'-triphosphate (ATP) release from postganglionic sympathetic nerves in rat small mesenteric arteries in vitro have been investigated on an impulse-by-impulse basis. NA release was measured using continuous amperometry and ATP release was monitored by intracellular recording of excitatory junction potentials (e.j.ps). 2. Electrical stimuli evoked transient increases in oxidation current. During trains of ten stimuli at 0.5 - 4 Hz there was a depression in the amplitude of oxidation currents evoked following the first stimulus in the train. 3. The neuronal NA uptake inhibitor, desmethylimipramine (1 microM), increased the amplitude of the summed oxidation current evoked by ten stimuli at 1 Hz and slowed the decay of oxidation currents evoked by trains of ten stimuli at 1 and 10 Hz. 4. The alpha2-adrenoceptor antagonist, idazoxan (1 microM), increased the amplitudes of the oxidation currents evoked during trains of ten stimuli at 0.5 - 10 Hz but had no effect on the oxidation currents evoked by the first stimulus in the train. 5. Idazoxan (1 microM) increased the amplitude of all e.j.ps evoked during trains of stimuli at 0.5 and 1 Hz. In addition, the facilitatory effect of idazoxan on e.j.ps was significantly greater than that on oxidation currents. 6. The findings indicate that NA release from sympathetic nerves supplying small mesenteric arteries is regulated by activation of presynaptic alpha2-adrenoceptors and that clearance of released NA in this tissue depends, in part, upon neuronal uptake. The different effects of idazoxan on the oxidation currents and e.j.ps may indicate that the release of NA and ATP is differentially modulated.
Subject(s)
Mesenteric Arteries/innervation , Norepinephrine/metabolism , Sympathetic Nervous System/metabolism , Adenosine Triphosphate/metabolism , Adrenergic Uptake Inhibitors/pharmacology , Adrenergic alpha-2 Receptor Antagonists , Adrenergic alpha-Antagonists/pharmacology , Animals , Cadmium/pharmacology , Corticosterone/pharmacology , Desipramine/pharmacology , Electric Conductivity , Electric Stimulation , Electrophysiology , Excitatory Postsynaptic Potentials/drug effects , Female , Idazoxan/pharmacology , In Vitro Techniques , Kinetics , Membrane Potentials/drug effects , Mesenteric Arteries/drug effects , Rats , Rats, Wistar , Receptors, Adrenergic, alpha-2/metabolism , Sympathetic Nervous System/drug effects , Tetrodotoxin/pharmacologyABSTRACT
1. Intracellularly recorded excitatory junction potentials (ej.ps) were used to study the effects of adenosine receptor antagonists on neurotransmitter release from postganglionic sympathetic nerve terminals in the guinea-pig vas deferens in vitro. 2. The A1 adenosine receptor antagonists, 8-phenyltheophylline (10 microM) and 8-cyclopentyl-1,3-dipropylxanthine (0.1 microM), increased the amplitude of e.j.ps evoked during trains of 20 stimuli at 1 Hz in the presence, but not in the absence, of the alpha2-adrenoceptor antagonist, yohimbine (1 microM) or the non-selective alpha-adrenoceptor antagonist, phentolamine (1 microM). 3. Adenosine (100 microM) reduced the amplitude of e.j.ps, both in the presence and in the absence of phentolamine (1 microM). This inhibitory effect of adenosine is most likely caused by a reduction in transmitter release as there was no detectable change in spontaneous ej.p. amplitudes. 4. In the presence of phentolamine, application of the adenosine uptake inhibitor, S-(p-nitrobenzyl)-6-thioinosine (0.1 microM), had no effect on ej.p. amplitudes. 5. The phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (100 microM), significantly increased the amplitudes of all e.j.ps evoked during trains of 20 stimuli at 1 Hz, both in the presence and in the absence of phentolamine (1 microM). 6. These results suggest that endogenous adenosine modulates neurotransmitter release by an action at prejunctional A1 adenosine receptors only when alpha2-adrenoceptors are blocked.
Subject(s)
Adenosine Triphosphate/metabolism , Purinergic P1 Receptor Antagonists , Synaptic Transmission/physiology , Vas Deferens/drug effects , 1-Methyl-3-isobutylxanthine/pharmacology , Adenosine/pharmacology , Adenosine/physiology , Adrenergic alpha-Antagonists/pharmacology , Animals , Guinea Pigs , In Vitro Techniques , Male , Membrane Potentials/drug effects , Membrane Potentials/physiology , Neuroeffector Junction/drug effects , Neuroeffector Junction/physiology , Phentolamine/pharmacology , Phosphodiesterase Inhibitors/pharmacology , Receptors, Purinergic P1/physiology , Synaptic Transmission/drug effects , Theophylline/analogs & derivatives , Theophylline/pharmacology , Thioinosine/analogs & derivatives , Thioinosine/pharmacology , Vas Deferens/innervation , Vas Deferens/physiology , Xanthines/pharmacology , Yohimbine/pharmacologyABSTRACT
1. The effects of Ca2+ concentration and Ca2+ channel blockers on noradrenaline (NA) and adenosine 5'-triphosphate (ATP) release from postganglionic sympathetic nerves have been investigated in rat tail arteries in vitro. Intracellularly recorded excitatory junction potentials (e.j.ps) were used as a measure of ATP release and continuous amperometry was used to measure NA release. 2. Varying the extracellular Ca2+ concentration similarly affected the amplitudes of e.j.ps and NA-induced oxidation currents evoked by trains of ten stimuli at 1 Hz. 3. The N-type Ca2+ blocker, omega-conotoxin GVIA (omega-CTX GVIA, 0.1 microM) reduced the amplitudes of both e.j.ps (evoked by trains of ten stimuli at 1 Hz) and NA-induced oxidation currents (evoked by trains of ten stimuli at 1 Hz and 50 stimuli at 10 Hz) by about 90%. 4. The omega-CTX GVIA resistant e.j.ps and NA-induced oxidation currents evoked by trains of 50 stimuli at 10 Hz were abolished by the non-selective Ca2+ channel blocker, Cd2+ (0.1 mM), and were reduced by omega-conotoxin MVIIC (0.5 microM) and omega-agatoxin IVA (40 nM). 5. Nifedipine (10 microm) had no inhibitory effect on omega-CTX GVIA resistant e.j.ps and NA-induced oxidation currents. 6. Thus both varying Ca2+ concentration and applying Ca2+ channel blockers results in similar effects on NA and ATP release from postganglionic sympathetic nerves. These findings are consistent with the hypothesis that NA and ATP are co-released together from the sympathetic nerve terminals.
Subject(s)
Adenosine Triphosphate/metabolism , Arteries/drug effects , Calcium Channel Blockers/pharmacology , Calcium/pharmacology , Norepinephrine/metabolism , Synaptic Transmission/drug effects , omega-Conotoxins , Animals , Arteries/innervation , Arteries/metabolism , Calcium/metabolism , Dose-Response Relationship, Drug , Electrophysiology , Evoked Potentials/drug effects , Female , In Vitro Techniques , Membrane Potentials/drug effects , Nifedipine/pharmacology , Peptides/pharmacology , Rats , Rats, Wistar , Sympathetic Nervous System/drug effects , Sympathetic Nervous System/metabolism , Tail , omega-Conotoxin GVIAABSTRACT
1. Extracellular recording techniques have been used to study nerve impulses in single sensory nerve terminals in guinea-pig cornea isolated in vitro. 2. Nerve impulses occurred spontaneously and were evoked by electrical stimulation of the ciliary nerves. 3. The nerve impulses were identified as originating in polymodal receptors, mechano-receptors or 'cold' receptors. All three types are believed to be nociceptors. 4. Tetrodotoxin (TTX, 1 microM) blocked nerve impulses evoked by electrical stimulation of the ciliary nerves. However, ongoing and/or naturally evoked nerve impulses persisted in the presence of TTX in all three types of receptors. Lignocaine (lidocaine; 1 mM) blocked all electrical activity. 5. TTX-resistant sodium channels therefore play a major role in generating the action potentials that signal pain to the brain.
Subject(s)
Cornea/innervation , Evoked Potentials/physiology , Nerve Endings/physiology , Nociceptors/physiology , Sodium Channels/physiology , Animals , Capsaicin/pharmacology , Ciliary Body/innervation , Electric Stimulation , Evoked Potentials/drug effects , Guinea Pigs , In Vitro Techniques , Lidocaine/pharmacology , Reaction Time , Sodium Channels/drug effects , Tetrodotoxin/pharmacologyABSTRACT
1. The effects of continuous but asynchronous nerve activity induced by ciguatoxin (CTX-1) on the membrane potential and contraction of smooth muscle cells have been investigated in rat proximal tail arteries isolated in vitro. These effects have been compared with those produced by the continuous application of phenylephrine (PE). 2. CTX-1 (0.4 nM) and PE (10 microM) produced a maintained depolarization of the arterial smooth muscle that was almost completely blocked by alpha-adrenoceptor blockade. In both cases, the depolarization was more sensitive to the selective alpha-adrenoceptor antagonist, idazoxan (0.1 microM), than to the selective alpha 1-adrenoceptor antagonist, prazosin (0.01 microM). 3. In contrast, the maintained contraction of the tail artery induced by CTX-1 (0.2 nM) and PE (2 and 10 microM) was more sensitive to prazosin (0.01) microM, than to idazoxan (0.01 microM). In combination, these antagonists almost completely inhibited contraction to both agents. 4. Application of the calcium channel antagonist, nifedipine (1 microM), had no effect on the depolarization induced by either CTX-1 or PE but maximally reduced the force of the maintained contraction to both agents by about 50%. 5. We conclude that the constriction of the tail artery induced by CTX-1, which mimics the natural discharge of postganglionic perivascular axons, is due almost entirely to alpha-adrenoceptor activation. The results indicate that neuronally released noradrenaline activates more than one alpha-adrenoceptor subtype. The depolarization is dependent primarily on alpha 2-adrenoceptor activation whereas the contraction is dependent primarily on alpha 1-adrenoceptor activation. The links between alpha-adrenoceptor activation and the voltage-dependent and voltage-independent mechanisms that deliver Ca2+ to the contractile apparatus appear to be complex.
Subject(s)
Arteries/physiology , Receptors, Adrenergic, alpha/physiology , Tail/blood supply , Adrenergic alpha-Antagonists/pharmacology , Animals , Arteries/drug effects , Arteries/innervation , Calcium Channel Blockers/pharmacology , Ciguatoxins/pharmacology , Female , Idazoxan/pharmacology , Membrane Potentials/drug effects , Muscle Contraction/drug effects , Nifedipine/pharmacology , Phenylephrine/pharmacology , Prazosin/pharmacology , Rats , Receptors, Adrenergic, alpha/drug effectsABSTRACT
1. Intraluminally perfused lymphatic vessels from the mesentery of the guinea-pig were examined in vitro to investigate their contractile activity and the co-ordination of this activity between adjacent lymphangions. 2. Lymphangions constricted at fairly regular intervals and exhibited 'refractory' periods of up to 3 s during which constrictions did not occur. 3. The contractile activity of adjacent lymphangions was highly co-ordinated. 4. The smooth muscle was found to be continuous between the adjacent lymphangions for the majority of valve regions examined morphologically (52 of 63 preparations). 5. Mechanical and electrical coupling between adjacent lymphangions was indicated, as some lymphangions underwent transient dilatations just prior to constriction, whereas direct electrophysiological measurements showed that the smooth muscle of most adjacent lymphangions was electrically coupled across the valve (15 out of 20 pairs of lymphangions). 6. It is concluded that perfused lymphangions of guinea-pig mesenteric lymphatic vessels rhythmically constrict, with the contractile activity of adjacent lymphangions highly co-ordinated. The findings also indicate that transmission of both mechanical and electrical signals between the adjacent lymphangions contribute to the co-ordination of their contractile activity.
Subject(s)
Lymphatic System/physiology , Muscle Contraction , Muscle, Smooth/physiology , Animals , Electrophysiology , Female , Guinea Pigs , In Vitro Techniques , Lymphatic System/cytology , Male , Membrane Potentials , Mesentery , Signal Transduction , Time FactorsABSTRACT
1. The effects of prejunctional beta-adrenoceptor activation on electrically evoked noradrenaline (NA) and adenosine 5'-triphosphate (ATP) were studied by use of continuous amperometry and conventional intracellular recording techniques. Excitatory junction potentials (e.j.ps) were used as a measure of ATP release, and NA-induced slow depolarizations and oxidation currents as measures of NA release, from postganglionic sympathetic nerves innervating the rat tail artery in vitro. 2. Isoprenaline (0.1 microM) increased the amplitude of e.j.ps, slow depolarizations and oxidation currents evoked by short trains of stimuli at 1 to 4 Hz. The facilitatory effect of isoprenaline on e.j.ps and oxidation currents was most pronounced on responses evoked by the first stimulus in a train. 3. Isoprenaline (0.1 microM) did not detectably alter the amplitude-frequency distribution of spontaneous e.j.ps. 4. The facilitatory effect of isoprenaline on e.j.ps, slow depolarizations and oxidation currents was abolished by the beta-adrenoceptor antagonist, propranolol (0.1 microM). Propranolol alone had no effect on e.j.ps, slow depolarizations or oxidation currents. 5. Thus, activation of prejunctional beta-adrenoceptors increases the release of both NA and ATP from postganglionic sympathetic nerves. The findings are consistent with the hypothesis that NA and ATP are released from the same population of nerve terminals and presumably from the same vesicles.
Subject(s)
Adenosine Triphosphate/metabolism , Arteries/innervation , Norepinephrine/metabolism , Receptors, Adrenergic, beta/physiology , Sympathetic Nervous System/metabolism , Tail/blood supply , Adrenergic beta-Agonists/pharmacology , Animals , Arteries/physiology , Electrochemistry , Isoproterenol/pharmacology , Male , Membrane Potentials/drug effects , Propranolol/pharmacology , Rats , Sympathetic Nervous System/physiology , Sympathomimetics/pharmacologyABSTRACT
An in vitro quantal assay (TCID50) for a non-occluded baculo-like virus isolate from naturally infected Penaeus japonicus obtained from China and experimentally infected P. stylirostris was developed using primary shrimp lymphoid cell cultures in Primaria 24-well tissue culture plates. The virus caused cytopathogenic effect (CPE) in the cell cultures as early as 2 day post-infection (p.i.). Initially, the cells rounded up and finally detached from the culture vessel as the infection progressed. At the present time, there is no established quantitative in vitro cell culture protocol for the assay of this baculo-like virus which has been reported by our laboratory to be highly pathogenic for P. stylirostris and P. vannamei, the two species of penaeid shrimp commercially cultured in Hawaii and the Western hemisphere. This quantal assay thus provides a simple and convenient method for the detection and assay of infectious virus in cultured penaeid shrimp.
Subject(s)
Baculoviridae/isolation & purification , Decapoda/virology , Animals , Cells, Cultured , Lymphocytes/cytology , Lymphocytes/virologyABSTRACT
It has previously been reported that the increase in blood pressure in the spontaneously hypertensive rat (SHR) occurs concurrently with a marked increase in thickness of the arterial wall and an increase in vascular innervation, particularly for the small muscular arteries. The purpose of the present study was to determine whether prevention of the increase in vascular innervation could prevent elevation of blood pressure in the SHR. We found that intraperitoneal injection of a single dose of an antiserum to nerve growth factor (anti-NGF) into young SHRs (postnatal day 19-24) caused a marked reduction in mean blood pressure at age 3-4 months from the raised value of 24.2 +/- 0.5 kPa to 18.9 +/- 0.8 kPa. By comparison, treated Wistar Kyoto rats (WKYs) maintained normal blood pressures. The treatment reduced the amplitude of the intracellularly recorded excitatory junction potential and the NA content of mesenteric arteries in the SHR, leaving the values similar to those of control WKYs. The NA content of these vessels was also reduced in treated WKYs. Importantly, the thickness of the vessel wall, which was greater in the SHR than the WKY, was not significantly altered by anti-NGF treatment. It is concluded that anti-NGF treatment during late neonatal development inhibits the increase in the functional levels of vascular innervation observed in the SHR. Furthermore, this increase in the functional levels of vascular innervation is necessary for the development of hypertension in this rat strain.
Subject(s)
Hypertension/prevention & control , Sympathectomy , Sympathetic Nervous System/physiology , Animals , Antibodies/pharmacology , Blood Pressure/physiology , Electrophysiology , Hypertension/physiopathology , Mesenteric Arteries/chemistry , Mesenteric Arteries/innervation , Mesenteric Arteries/physiopathology , Nerve Growth Factors/immunology , Norepinephrine/metabolism , Rats , Rats, Inbred SHR , Rats, Inbred WKY , Vasoconstrictor Agents/metabolismABSTRACT
Fluorescence in situ hybridization was used in interphase cells of 30 ovarian carcinomas to detect numerical changes in copy number of 13 different centromeres (1, 2, 3, 4, 6, 7, 8, 10, 11, 12, 17, 18, and X). Thirty-seven percent of samples (11/30) were near diploid and demonstrated only minor changes in centromere copy number, involving gain and/or loss of one or a few centromeres. The most common changes included loss of centromeres 4,6, 17, and 18 and gain of centromere 1. The remaining 63% of samples were hyperdiploid and demonstrated a general increase in copy number of most or all centromeres examined. Among these samples, the centromere of chromosome I was most often found to be at higher copy number. Centromeres that were less often at increased copy or deleted within the hyperdiploid samples include centromeres 4, 17, 18, and X. These results suggest that tumor-suppressor genes that are located on chromosomes 4, 6, 17, and 18 may be involved in the development and progression of ovarian cancer.