Life-long learning in laboratory animal science and ethics for veterinary and para-veterinary professionals in South Africa.

Veterinary and para-veterinary professionals working in the animal research sector are critical to ensure scientific quality and the humane care and use of animals. However, there are few focused education and training opportunities available for these professionals in South Africa. A survey of veterinarians working in animal research, undertaken by the South African Association for Laboratory Animal Science, identified the need for more advanced education and training opportunities beyond the routine Day 1 Skills currently provided for in undergraduate education. These could be broadly categorised into knowledge and skills relating to species-specific husbandry, procedures and clinical approaches, research-related biosecurity and biosafety, and study-specific ethical and animal welfare considerations. A subsequent workshop, attended by 85 veterinary and para-veterinary professionals in the animal research sector, identified 53 life-long learning needs, each with an associated learning outcome, for this professional community. These were grouped into five overarching themes: Personal development (9); Leadership and management skills (12); Education and training skills (5); Welfare, ethics and clinical skills (20); and Regulations and quality-assurance (7). Of the 53 learning outcomes, 14 were knowledge-based, ten were competencies, and 29 both knowledge and competence. These life-long learning opportunities, if available and implemented, will address important needs of veterinary and paraveterinary professionals in the animal research sector in South Africa. This would empower these professionals, assist in improving animal and human wellbeing, support high-quality ethical science, and maintain public confidence in the sector, thus enabling a more satisfactory career environment.

Learning outcomes for the education and training of laboratory animal caretakers in South Africa.

Education and training is essential for laboratory animal caretakers (LACs), but there are no courses available in South Africa. A national workshop was thus held to collaboratively establish the learning outcomes (LOs) for the education and training (E&T) of LACs. Eighty-five stakeholders from 30 institutions took part in small group discussions interspersed with plenary sessions to draw up the consensus LOs. One-hundred-and-twenty LOs were identified, grouped into the following three main themes and 15 topics: 1) Focus on animals (animal care and husbandry, animal ethics, animal welfare, basic biology, environment); 2) Focus on humans (administration, health and safety, lifelong learning, professionalism, psychological wellbeing); and 3) Focus on systems (biosecurity, equipment, jurisprudence, logistics, and quality management). This E&T framework provides a foundation for a career path in the laboratory animal science field. The psychological (i.e. mental and emotional) wellbeing of LACs forms a noteworthy component of the focus on humans, since working with research animals is stressful and coping mechanisms are needed in order to promote compassion satisfaction and prevent compassion fatigue and burnout. Approximately 75% of the LOs are knowledge-based, while 25% are competencies in practical skills. It is recommended that competencies should be assessed by direct observation of practical/procedural skills, where competence in a procedure or practical task is assessed against predetermined criteria. These LOs are published with the intent that they will promote animal and human wellbeing, support ethical science, maintain public confidence, and in so doing, contribute to a just and civilised society.

Review article: methods of measuring gastric acid secretion.

BACKGROUND: Gastric acid has an important pathophysiological role in human beings. Numerous methods have been evaluated over the years in an attempt to measure gastric acid and stomach acidity, to study the role of gastric acid in gastrointestinal diseases in humans and to evaluate the effects of acid suppressing drugs. AIM: To review methods that have been used to measure gastric acid and gastric acidity. METHODS: Searches of the electronic databases PUBMED, MEDLINE and EMBASE, were performed with articles restricted to English language and human subjects. References were also identified from the bibliographies of selected articles. RESULTS: Methods for measuring gastric acid include both invasive and non-invasive techniques. Invasive tests include the conventional gastric acid aspiration tests, gastric pH measurement techniques and endoscopic methods. Non-invasive methods use urinary analysis, breath analysis, serum pepsinogens assay, scintigraphic techniques, impedence tomography and alkaline tide for measurement of gastric acid. CONCLUSIONS: Several methods of measuring gastric acid exist. Invasive tube tests are uncomfortable and time consuming, whereas most of the non-invasive methods are at best semiquantitative and useful in detecting low or absent acid secretion. Further attempts to explore new methods for measuring gastric acid are therefore warranted.

Electrophysiological characteristics of vasomotor preganglionic neurons and related neurons in the thoracic spinal cord of the rat: an intracellular study in vivo.

Sympathetic preganglionic neurons (SPN) represent the final central neurons in the sympathetic pathways which regulate vasomotor tone; they therefore play a pivotal role in the re-distribution of cardiac output to different vascular beds in response to environmental challenges. While the consensus view is that activity in these neurons is due mainly to supraspinal inputs, the possibility that some activity may be generated intrinsically and modified by synaptic inputs cannot be excluded. Therefore, in order to distinguish between these two possibilities, the electrophysiological properties of cardiovascular-like SPN in the upper thoracic spinal cord of the anesthetized rat were examined and their response to activation of vasodepressor inputs was investigated. Intracellular recordings were made from 22 antidromically identified SPN of which 17 displayed irregular, but maintained, spontaneous activity; no evidence of bursting behavior or pacemaker-like activity was observed. Stimulation of the aortic depressor nerve or a vasodepressor site within the nucleus tractus solitarius (NTS) resulted in a membrane hyperpolarization, decrease in cell input resistance and long-lasting cessation of neuronal firing in SPN including a sub-population which had cardiac-modulated patterns of activity patterns. Recordings were also undertaken from 80 non-antidromically-activated neurons located in the vicinity of SPN; 23% of which fired in phase with the cardiac cycle, with this peak of activity occurring before similar increases in cardiac-modulated SPN. Stimulation of vasodepressor regions of the NTS evoked a membrane hyperpolarization and decrease in cell input resistance in cardiac-modulated but not non-modulated interneurons. These studies show that activity patterns in SPN in vivo are determined principally by synaptic inputs. They also demonstrate that spinal interneurons which exhibit cardiac-modulated patterns of activity are postsynaptically inhibited following activation of baroreceptor pathways. However, the question as to whether these inhibitory pathways and/or disfacilitation of tonic excitatory drive underlies the baroreceptor-mediated inhibition of SPN remains to be determined.

Enkephalinergic inhibition of raphe pallidus inputs to rat hypoglossal motoneurones in vitro.

Hypoglossal motoneurones play a major role in maintaining the patency of the upper airways and in determining airways resistance. These neurones receive inputs from many different regions of the neuroaxis including the caudal raphe nuclei. Whilst we have previously shown that glutamate is utilised in projections from one of these caudal raphe nuclei, the raphe pallidus, to hypoglossal motoneurones, these raphe pallidus-hypoglossal projections also contain multiple co-localised neuropeptides, including a population that are immunopositive for enkephalin. The role of enkephalin in the control of hypoglossal motoneurones is unknown. Therefore the aim of these studies was to determine whether enkephalins modulate caudal raphe glutamatergic inputs to hypoglossal motoneurones. Whole cell recordings were made from rat hypoglossal motoneurones in vitro, with glutamate-mediated excitatory postsynaptic currents (EPSCs) evoked in these neurones following electrical stimulation within the raphe pallidus. Superfusion of enkephalin significantly decreased the amplitude of these raphe pallidus evoked EPSCs (56.1+/-29% of control, P<0.001), an action that was mirrored by the tau-opioid receptor agonist, [D-Ala, N-Me-Phe, Gly-ol]-enkephalin acetate (DAMGO;53.8+/-26%, P<0.01), but not by the delta-opioid receptor agonist, [D-Pen]-enkephalin (DPDPE). Enkephalin also increased the amplitude ratio (1.57+/-0.36 vs. 1.14+/-0.27, P<0.01) of pairs of evoked EPSCs (paired pulse ratio), decreased the frequency (P<0.0001) but not the amplitude of miniature EPSCs, whilst having no effect on the inward current evoked by glutamate applied directly to the postsynaptic cell (97.8+/-2.2% of control, P=n.s.). Likewise, DAMGO also increased the paired pulse ratio (1.62+/-0.35 vs. 1.31+/-0.14, P<0.05) and decreased the frequency of miniature EPSCs (P<0.0001). Together, these data suggest that enkephalin acts at tau-opioid receptors located on the presynaptic terminals of raphe pallidus inputs to hypoglossal motoneurones to significantly decrease glutamate release from these projections.

Adrenaline modulates multiple conductances in rat vagal motoneurones in vitro.

Whole cell recordings were undertaken from vagal motoneurones, including identified gastric vagal motoneurones, located within the medial regions of the dorsal vagal motonucleus of the rat medulla in vitro. The actions of adrenaline on individual channels expressed by these neurones were investigated. Adrenaline directly inhibits Ca2+ currents and delayed rectifier K+ currents and activates a sustained Na+ current. It also inhibits both Ca2+ activated non-selective cationic currents and Ca2+ activated K+ currents, the latter via inhibition of the underlying activating Ca2+ current. Since different sub-populations of vagal motoneurones express different complements of ion channels, this selective modulation of specific conductances by adrenaline may provide a mechanism by which adrenergic inputs, which project non-selectively throughout the DMV, could selectively control different aspects of vagal function.

Ventral medullary chemoreceptor inputs to neurons within the nucleus ambiguus.

1. The neural mechanisms by which neurons within the nucleus ambiguus respond to chemoreceptor stimuli applied to the ventral medullary surface (VMS) were investigated by determining the effect of low pH on the membrane potential and synaptic activity of these neurons in vitro. 2. Small reductions in pH evoked an indirect membrane depolarization and/or an increase in excitatory synaptic activity in the majority of neurons. A direct membrane hyperpolarization was observed in the remaining neurons. 3. Acetylcholine evoked a direct nicotinic receptor-mediated membrane depolarization in these neurons. In addition, 37% of neurons received muscarinic synaptic input that originated from neurons located near the VMS. 4. Low-pH artificial cerebrospinal fluid (aCSF) potentiated the cholinergic component of the excitatory post-synaptic potential (EPSP) evoked from near the VMS. Both this EPSP and the spontaneous EPSP evoked by low-pH aCSF could be blocked by atropine. 5. It is concluded that at least two different mechanisms exist to transmit chemoreceptive information from the VMS to neurons within the nucleus ambiguus. In the majority of neurons, an indirect excitatory response is observed that is due, in part, to activation of a polysynaptic cholinergic pathway originating near the VMS. In the remaining neurons, low pH evokes a hyperpolarization that is due to a direct action of the dendrites of the neurons themselves that project near to the VMS.

Evidence that the firing pattern of sympathetic preganglionic neurones is determined by an interaction between amines and an excitatory amino acid.

Extracellular recordings were made from identified sympathetic preganglionic neurones (SPN) located in the second and third thoracic segments of the adult rat spinal cord, in-vivo. Iontophoretic application of the amines, 5-hydroxytryptamine (5-HT) and noradrenaline (NAdr) in the vicinity of these neurones evoked either long lasting excitations, inhibitions or bi-phasic changes (inhibition followed by excitation, 5-HT only) in neuronal firing rate. The excitatory response to both 5-HT or NAdr could only be obtained in spontaneously active neurones, or silent neurones, in which a subliminal level of an excitatory amino acid (EAA) was also present. In 4 neurones, the response evoked by NAdr was dependent on neuronal firing rate and hence the level of EAA present. At low basal firing rates, inhibitions were observed whereas at higher firing rates, excitations were observed. Subliminal levels of either 5-HT or NAdr markedly potentiated the increases in neuronal firing rate evoked by an EAA. In a sub-population of SPN, both 5-HT (18/88 neurones) and NAdr (21/51 neurones) induced a repeated bursting pattern of action potential discharge during the amine evoked changes in neuronal firing rate in otherwise irregularly discharging neurones. We conclude that these actions of the monoamines, that is excitations or inhibitions, gain enhancement, prolongation of action, burst firing and oscillations are likely to be due to the modulation by these agents of intrinsic membrane conductances. The differing responses evoked in SPN will produce marked differences in the efficacy of synaptic transmission in the sympathetic ganglia. As a result of these mechanisms, there will be a greater versatility in the sympathetic control systems than would otherwise exist.

Baroreceptor-induced inhibition of sympathetic neurons by GABA acting at a spinal site.

gamma-Aminobutyric acid (GABA) synapses are an important feature of sympathetic circuits in the spinal cord. The possibility that these spinal synapses participate in the reduction of sympathetic activity resulting from arterial baroreceptor activation was tested. For this purpose baroreceptors were stimulated by a rapid rise in blood pressure induced by intravenous phenylephrine, and the effect of this on a spinally evoked excitatory response in a renal sympathetic nerve was examined before, during, and after removal of tonic excitatory drive from the rostral ventrolateral medulla (RVLM). Stimulation of descending excitatory axons in the spinal cord at the fourth cervical spinal level evoked a biphasic excitatory response in a renal sympathetic nerve, with a latency of 56 +/- 6 ms and a duration of 230 +/- 24 ms. The magnitude of this spinally evoked response was significantly reduced (P < 0.001) during baroreceptor activation, thus confirming that there is a spinal component of the baroreceptor inhibitory reflex. Intrathecal administration of bicuculline reduced baroreceptor-mediated inhibition of the spinally evoked response to 48 +/- 18% of control inhibition (P < 0.05). Subsequent removal of tonic excitatory drive by microinjection of glycine into the RVLM produced no significant further change in the baroreceptor-mediated inhibition. This suggests that a GABAA receptor-mediated effect at a spinal site is involved in the baroreceptor-induced inhibition. Reversing this procedure by first placing injections of glycine into the RVLM resulted in a reduction of baroreceptor inhibition to 60 +/- 16% (P < 0.05) of control baroreceptor inhibition. Subsequent intrathecal administration of bicuculline produced no significant further change in the baroreceptor-mediated inhibition of the spinally evoked response. This suggests that the GABAA-mediated effect of the baroreceptors at a spinal site is dependent on the integrity of neurons in the RVLM. We conclude that GABA in sympathetic circuits in the spinal cord plays a significant role in the modification of sympathetic vasomotor activity by the baroreceptor reflex.

Bulbospinal catecholamine neurones and sympathetic pattern generation.

Populations of neurones containing noradrenaline, dopamine and possibly adrenaline project to the spinal cord where they innervate sympathetic preganglionic neurones (SPN). Studies are described which illustrate the actions of catecholamines on SPN and which suggest ways in which the catecholamine neurones could regulate the cardiovascular system. Experiments on rats using, intrathecal application of drugs to the spinal cord, iontophoresis, or superfusion of drugs whilst recording either postganglionic nerve activity, extracellularly or intracellularly from SPN respectively, reveal that catecholamines may excite or inhibit SPN. A slow depolarisation is mediated by alpha 1 adrenoceptors whereas alpha 2 adrenoceptors mediate a slow hyperpolarisation. Catecholamines may also excite glycinergic interneurones which elicit fast IPSPs in SPN. By regulating different ionic conductances in the membrane of SPN catecholamines are able to induce SPN to discharge tonically or to oscillate with bursts of action potentials. Furthermore these actions may be modified in the presence of an excitatory amino acid. It is suggested that via these mechanisms differential responses in the sympathetic outflow could be produced.

Chemical mediators of spinal inhibition of rat sympathetic neurones on stimulation in the nucleus tractus solitarii.

1. This study was undertaken to gain more direct evidence of the pathways and neurochemical mediators of a spinally mediated baroreceptor inhibition of sympathetic preganglionic neurones (SPNs). 2. For this purpose, single-pulse electrical stimulation within identified vasodepressor regions of the nucleus tractus solitarii (NTS) was used together with extracellular recordings of single antidromically identified SPNs in the T2 segment of the spinal cord of anaesthetized rats. 3. The actions of agonists and antagonists of inhibitory amino acids on the NTS-induced inhibitions were determined, when they were iontophoretically applied in the vicinity of SPNs via a multibarrel micropipette assembly. 4. Extracellular recordings were made from sixty-nine SPNs. In forty-four SPNs, NTS stimulation elicited a period of inhibition of activity in both spontaneous and 'D,L-homocysteic acid-driven' SPNs with a latency to onset of 60 +/- 6 ms and a magnitude of 80 +/- 3%. 5. In six out of eight neurones, the NTS-induced inhibition was reduced by 74 +/- 16% during the application of the glycine antagonist strychnine (0-10 nA, 5-10 min) with doses that selectively blocked the inhibitory effect of iontophoretically applied glycine. 6. In nine out of nine neurones, the NTS-induced inhibition was reduced by 38 +/- 6% during the application of the GABAA antagonist bicuculline (5-15 nA, 4-14 min) with doses that selectively blocked the inhibitory effect of iontophoretically applied GABA. 7. In two SPNs, the actions of strychnine and bicuculline were shown to be additive in blocking the NTS inhibition. 8. The selective GABAB antagonists, CGP 35348 (20-50 nA, 6-25 min) and CGP 55845A (10 nA, 11 min) did not antagonize the NTS-induced inhibition. 9. It is suggested that GABA and glycine interneurones are activated by a baroreceptor bulbospinal pathway to inhibit sympathetic preganglionic neurones in the spinal cord.

The spinal organisation of the baroreceptor reflex.

Baroreceptor inhibition of a spinally evoked response in a renal nerve was studied following removal of excitatory drive from the rostral ventrolateral medulla (RVLM), by microinjecting glycine into this region (RVLM block). Activation of arterial baroreceptors was still able to inhibit a spinally evoked response after RVLM block and this effect was abolished by intrathecal strychnine. Intrathecal bicuculline also was shown to reduce the magnitude of the baroreceptor inhibition but only when the RVLM was intact indicating that bicuculline was removing a facilitation. Both strychnine and bicuculline antagonised an NTS induced inhibition of activity in single sympathetic preganglionic neurones. It is concluded that arterial baroreceptor reflex regulation of vasomotor activity occurs at a spinal as well as a supraspinal site and GABA and glycine are the likely inhibitory mediators at both sites.

Mediation of baroreceptor inhibition of sympathetic nerve activity via both a brainstem and spinal site in rats.

1. The possible involvement of a spinal site of sympatho-inhibitory action in the baroreceptor reflex pathways was investigated by determining the effect of phenylephrine-induced, baroreceptor-mediated inhibition on a spinally evoked excitatory response of renal nerve activity before, during and after removal of tonic descending excitatory drive from the rostral ventrolateral medulla (RVLM). 2. Stimulation of descending excitatory axons at the C4 vertebral level evoked a highly reproducible excitatory response (mean variance, 2.2 +/- 0.6%) in a renal sympathetic nerve with a latency of 72 +/- 5.7 ms and duration of 211 +/- 32 ms. 3. Following baroreceptor activation, the magnitude of this spinally evoked response was reduced by 33.5 +/- 4.2% compared with control spinal response. 4. To remove tonic descending excitatory drive, glycine was microinjected bilaterally into the RVLM (RVLM block); this briefly and reversibly abolished spontaneous sympathetic nerve activity. 5. Following RVLM block, the spinally evoked response in a renal sympathetic nerve was inhibited by 16.9 +/- 4.4% during baroreceptor activation. 6. Intrathecal administration of the glycine antagonist strychnine to the lower thoracic segments of the spinal cord virtually abolished this inhibition. 7. It was concluded that baroreceptor inhibition of sympathetic activity occurs at a spinal site as well as a supraspinal one. Glycine is a likely mediator of the inhibition at the spinal site.

Dye-coupling between vagal motoneurones within the compact region of the adult rat nucleus ambiguus, in-vitro.

The electrophysiological and morphological characteristics of vagal motoneurones lying within the compact region of the nucleus ambiguus were investigated in thin coronal slices of the adult rat medulla utilising intracellular recording techniques. The majority of neurones were found to be silent, displaying no underlying synaptic activity or oscillations in membrane potential. Intracellular dye-filling demonstrated that the neurones had multipolar cell bodies, with 2-8 major dendrites, each branching up to 4 times and extending up to 200 microns from the cell body. The existence of dye-coupling between adjacent neurones was shown in 30% of cells investigated. This evidence suggests a possible mechanism for the provision of synchronous activity within groups of vagal motoneurones, a process essential for the control of deglutination.

Angiotensin II in the spinal cord of the rat and its sympatho-excitatory effects.

Immunohistochemical studies have shown there is a dense angiotensin-like immunoreactivity of terminals in the sympathetic region of the thoracic and lumbar spinal cord. In the present study measurements were made of the concentration of angiotensin in the spinal cord of rats using radioimmunoassay following two different extraction procedures. These gave concentrations of angiotensin as mean of 108 and 161 pg.g-1 tissue wet weight. Angiotensin II given intrathecally or microinjected into the spinal cord caused an increase in postganglionic sympathetic nerve activity which was blocked by prior application of saralasin. Angiotensin III was without effect. Intracellular recordings from sympathetic preganglionic neurones in-vitro in slices of neonate rat spinal cord showed that angiotensin II produced an increase of excitability of the neurones by a slow depolarisation without the generation of action potentials. This effect still occurred in the presence of TTX. Angiotensin II also could increase synaptic activity, both EPSPs and IPSPs as well as a synaptically induced slow depolarisation being observed suggesting that presympathetic interneurones are also sensitive to the peptide. The evidence indicates that if angiotensin is released from nerve terminals surrounding sympathetic neurones it will enhance the gain of the neurone so that it could more easily be discharged by other excitatory inputs.

Excitatory and indirect inhibitory actions of 5-hydroxytryptamine on sympathetic preganglionic neurones in the neonate rat spinal cord in vitro.

The action of 5-hydroxytryptamine (5-HT) on sympathetic preganglionic neurones (SPN) was studied by intracellular recordings in thin slices of neonatal rat spinal cord in vitro. Superfusion of 5-HT (1-270 microM) to SPN caused a concentration dependent slow depolarization or inward current and an increase in synaptic activity consisting of both EPSPs and IPSPs. The slow depolarization was still present after superfusion with TTX. Similar effects were seen during superfusion with 5-carboxamidotryptamine (5-CT) or alpha-methyl-5-hydroxytryptamine (alpha-me-5-HT). A comparison with the potency of 5-HT was made for 5-CT or alpha-me-5-HT on the same neurone by determining the magnitude of the slow depolarization to different concentrations of agonist. This showed that the apparent potency of the agonists was 5-CT > 5-HT > alpha-me-5-HT even in the presence of fluoxetine, a 5-HT uptake inhibitor. The 5-HT-induced slow depolarization was partially blocked by ketanserin but full recovery was not observed. The results suggest that the excitatory action of 5-HT on SPN is mediated via an atypical 5-HT2 receptor or a 5-HT1C-like receptor. The 5-HT-induced IPSPs were reversibly blocked by superfusion with strychnine, suggesting they were mediated by glycine.

The actions of 5-hydroxytryptamine on the membrane of putative sympatho-excitatory neurones in the rostral ventrolateral medulla of the adult rat in vitro.

Intracellular recordings were obtained in vitro from neurones lying within the rostral ventrolateral medulla of the adult rat. Neurones could be classified into three groups: silent neurones, irregularly firing neurones which had a regular pattern of action-potential generation. Membrane hyperpolarization of regularly firing neurones failed to reveal underlying EPSPs or disrupt the regular pattern of action-potential generation. Superfusion of a high Mg2+, low Ca2+ aCSF did not abolish action-potential generation but the regular pattern of firing of these neurones was lost. 5-Hydroxytryptamine evoked a slow concentration-dependent hyperpolarization in both spontaneously active and silent neurones, accompanied by a decrease in cell-input resistance. This study has provided further evidence for pacemaker-like neurones within the RVLM and for the modulation of these neurones by 5-hydroxytryptamine.

Excitation and inhibition of rat sympathetic preganglionic neurones by catecholamines.

The actions of microiontophoretically applied catecholamines on antidromically identified sympathetic preganglionic neurones (SPN) in the upper thoracic spinal cord of the anaesthetized rat were investigated. Noradrenaline (NA) excited the majority of neurones (50/71), however, a significant number were inhibited by the catecholamine (17/71). Adrenaline excited 4/9 SPN and inhibited 2/9. Dopamine had excitatory actions on SPN (3/3). Dual actions of NA on the same SPN were demonstrated, with the actions of the catecholamine being modulated by excitatory amino acids. NA was also shown to induce burst firing in 21% of SPN.

The influence of 5-hydroxytryptamine agonists and antagonists on identified sympathetic preganglionic neurones in the rat, in vivo.

1. 5-Hydroxytryptamine (5-HT) was applied by microiontophoresis in the vicinity of identified sympathetic preganglionic neurones in the upper thoracic spinal cord of the rat, in vivo. 2. Sympathetic preganglionic neurones responded in one of three ways to 5-HT: by (a) excitation (76%), (b) inhibition (4%) or (c) in a biphasic manner (5%). 3. The excitatory responses evoked by 5-HT were mimicked by 5-carboxamidotryptamine (5-CT) and alpha-methyl-5-hydroxytryptamine (alpha-Me-5-HT). The inhibitory and biphasic responses evoked by 5-HT were mimicked by 2-methyl-5-hydroxytryptamine (2-Me-5-HT). The observed responses evoked by 5-HT and selective agonists may be different on the same cell. In several instances a single neurone excited by one agonist was inhibited by another agonist. 4. The 5-HT2-receptor antagonists, ketanserin and LY 53857, failed to abolish selectively the excitatory responses evoked by 5-HT and alpha-Me-5-HT, when applied by microiontophoresis. The antagonists non-selectively reduced the excitatory responses evoked by 5-HT, 5-CT, alpha-Me-5-HT, D,L-homocysteic acid (DLH) and noradrenaline (NA). A reduction in synaptically evoked activity was also observed. 5. The 5-HT3-receptor antagonist, ICS 205-930, failed to abolish the inhibitory responses evoked by 5-HT. 6. It was concluded that the excitatory responses evoked by 5-HT are mediated by a receptor that is neither 5-HT2 or 5-HT3, but shows similarities to the 5-HT1-like receptor profile. The inhibitory actions of 5-HT are mimicked by 2-Me-5-HT, but the receptor is not 5-HT3, or 5-HT1-like or 5-HT2.

Effects of activating spinal alpha-adrenoreceptors on sympathetic nerve activity in the rat.

The effects of several alpha-adrenoreceptor agonists and antagonists administered intrathecally at T10 level on renal sympathetic nerve activity (RSNA) were examined, in chloralose-urethane anaesthetised rats. Intrathecal noradrenaline (NA, 0.5-500 micrograms) produced one of 3 responses depending on dose, an inhibition of RSNA at low doses, an excitation of RSNA at high doses, or a biphasic effect. Intrathecal adrenaline (5-200 micrograms) was inhibitory in the main but some doses elicited poorly repeatable brief excitation followed by prolonged inhibition. Intrathecal methoxamine (ME; 2.5 ng-25 micrograms) caused a dose-dependent increase in RSNA (mean maximum response 27 +/- 0.5%). The excitatory effects of NA and ME were blocked (72% +/- 12%) by pretreatment with intrathecal prazosin (PRA, 20-200 ng) but not by yohimbine (YOH, 200 ng). Intrathecal guanabenz (GUA 3-15 micrograms) caused a dose-dependent inhibitory effect on RSNA (mean maximum 32% +/- 5%). The inhibitory effects of NA, adrenaline and GUA were blocked by pretreatment with intrathecal YOH (200 ng-2 micrograms). Intrathecal PRA (200 ng) had no effect on the inhibitory effects of NA and GUA. Intravenous administration of each of the adrenoreceptor agonists (apart from adrenaline), at similar doses to those given intrathecally, in most cases had no significant effect on RSNA; in a few cases the opposite effects to those produced by intrathecal administration were seen.