Regina Fritsche

Autonomic nervous control of blood pressure and heart rate during hypoxia in the cod, Gadus morhua

SummaryThe autonomic nervous and possible adrenergic humoral control of blood pressure and heart rate during hypoxia was investigated in Atlantic cod. The oxygen tension in the water was reduced to 4.0–5.3 kPa (i.e.. PwO2=30–40 mmHg), and the fish responded with an immediate increase in ventral and dorsal aortic blood pressure (PvaPda), as well as a slowly developing bradycardia. The plasma concentrations of circulating catecholamines increased during hypoxia with a peak in the plasma level of noradrenaline occurring before the peak for adrenaline. Bretylium was used as a chemical tool to differentiate between neuronal and humoral adrenergic control of blood pressure and heart rate (fH) during hypoxia. The increase in Pva and Pda in response to hypoxia was strongly reduced in bretylium-treated cod, which suggests that adrenergic nerves are responsible for hypoxic hypertension. In addition, a small contribution by circulating catecholamines to the adrenergic tonus affecting Pva during hypoxia was suggested by the decrease in Pva induced by injection of the α-adrenoceptor antagonist phentolamine. The cholinergic and the adrenergic tonus affecting heart rate were estimated by injections of atropine and the β-adrenoceptor antagonist sotalol. The experiments demonstrate an increased cholicholinergic as well as adrenergic tonus on the heart during hypoxia.

Cardiovascular and ventilatory control during hypoxia

The cardiovascular and respiratory physiology of fish has been an area of major interest in comparative physiology for several decades, but the systems that control these functions and the nature of the nervous reflexes that regulate cardiovascular and respiratory functions have received relatively little interest. Research on cardiovascular control systems has been largely restricted to adrenergic vasomotor control - adrenergic neurones and circulating catecholamines - and of course also the double antagonistic cholinergic and adrenergic innervation of the teleost heart. In addition, recent studies have discussed whether catecholamines are involved in ventilatory control in teleosts (Aota et al., 1990; Playl et al., 1990; Kinkead and Perry, 1990,1991; Perry et al., 1991).

Respiratory and cardiovascular responses to hypoxia in the Australian lungfish.

Simultaneous measurements of pulmonary blood flow (qPA), coeliacomesenteric blood flow (qCoA), dorsal aortic blood pressure (PDA), heart rate (fH) and branchial ventilation frequency (fv) were made in the Australian lungfish, Neoceratodus forsteri, during air breathing and aquatic hypoxia. The cholinergic and adrenergic influences on the cardiovascular system were investigated during normoxia using pharmacological agents, and the presence of catecholamines and serotonin in different tissues was investigated using histochemistry. Air breathing rarely occurred during normoxia but when it did, it was always associated with increased pulmonary blood flow. The pulmonary vasculature is influenced by both a cholinergic and adrenergic tonus whereas the coeliacomesenteric vasculature is influenced by a beta-adrenergic vasodilator mechanism. No adrenergic nerve fibers could be demonstrated in Neoceratodus but catecholamine-containing endothelial cells were found in the atrium of the heart. In addition, serotonin-immunoreactive cells were demonstrated in the pulmonary epithelium. The most prominent response to aquatic hypoxia was an increase in gill breathing frequency followed by an increased number of air breaths together with increased pulmonary blood flow. It is clear from the present investigation that Neoceratodus is able to match cardiovascular performance to meet the changes in respiration during hypoxia.

Immunohistochemical localization of bioactive peptides and amines associated with the chromaffin tissue of five species of fish.

Biogenic peptides and amines associated with the chromaffin tissue in Atlantic cod (Gadus morhua), rainbow trout (Oncorhynchus mykiss), European eel (Anguilla anguilla), spiny dogfish (Squalus acanthias) and Atlantic hagfish (Myxine glutinosa) were identified utilizing immunohistochemical techniques. Within the posterior cardinal vein (PCV) in cod, trout and eel, a subpopulation of chromaffin cells displayed immunoreactivity to tyrosine hydroxylase (TH) and dopamine-β-hydroxylase (DβH) but not to phenylethanolamine-N-methyltransferase (PNMT). TH-like immunorectivity was observed within cells in hagfish hearts. Nerve fibres displaying vasoactive intestinal peptide (VIP) immunoreactivity and pituitary adenylyl cyclase activating peptide (PACAP) immunoreactivity innervated cod, trout and ell chromaffin cells. In eel, neuropeptide Y (NPY)-like and peptide YY (PYY)-like immunoreactivity was located within cells in the PCV, including chromaffin cells. Serotonin-like immunoreactivity was observed within eel and cod chromaffin cells and in hagfish hearts. In the dogfish axillary bodies, nerves displaying TH-like, VIP-like, PACAP-like, substance P-like and galanin-like immunoreactivity were observed. These results are compared with those of other vertebrates, and potential roles for these substances in the control of catecholamine release are suggested.

Understanding cardiovascular physiology in zebrafish and Xenopus larvae: the use of microtechniques

Zebrafish and Xenopus, genetically accessible vertebrates with an externally developing, optically clear embryo, are ideally suited for in vivo functional dissection of the embryonic development of the circulatory system. Physiological characterizations of the cardiovascular system are still imperative for a more complete understanding of the connections between genetic/epigenetic factors and cardiovascular development. Here, we review experimental tools and methods that have been developed to measure numerous cardiovascular parameters in these millimetre-sized animals.

The Role of Circulating Catecholamines in the Ventilatory and Hypertensive Responses to Hypoxia in the Atlantic Cod (Gadus morhua)

This is the first description of the ventilatory response of the Atlantic cod (Gadus morhua) to hypoxia. Using a pharmacological approach, we tested the hypotheses that during hypoxia (1) elevation of circulating catecholamines, arising from chromaffin tissue or peripheral adrenergic neurones, contributes to hyperventilation and that (2) the hyperventilatory response can be modulated by the hypoxia-induced hypertension. The ventilatory response to hypoxia (final water PO₂ = 46 mmHg, reached in ≤25 min) was not significantly altered by pretreatment offish with either α-or β-adrenoceptor antagonists (phentolamine or sotalol, respectively) or with an inhibitor of catecholamine release from peripheral neurones (bretylium), despite a significant increase in plasma catecholamine levels in all treatment groups. Since the utilized regime did not induce metabolic acidosis (a potential ventilatory stimulant), we conclude that the hyperventilation was caused solely by a depression of the oxygen status. Although changes in the internal as well as the external oxygen status may have potentially contributed to the hyperventilation, we suggest, on the basis of the rapid ventilatory response after only small depressions of water PO₂, that the initial stimulus is external. During hypoxia, a doubling of mean ventral aortic blood pressure was observed. This elevation of blood pressure was due to an increased adrenergic nervous and humoral activity. The lack of modification in the ventilatory response to hypoxia after transient or persistent abolishment of the hypertension by pharmacological agents (bretylium or phentolamine, respectively) indicated that during hypoxia there was no relationship between blood pressure and ventilation in this species.

Neuropeptide immunoreactivity and co-existence in cardiovascular nerves and autonomic ganglia of the estuarine crocodile, Crocodylus porosus, and cardiovascular effects of neuropeptides

The two aortas of the crocodile are in open connection at two sites, the foramen of Panizzae immediately outside the ventricles, and the arterial anastomosis at the level of the gut. The present study was performed to elucidate the innervation of the cardiovascular structures of the crocodile, in part to provide a further basis for the assumption that the apertures of the foramen and the anastomosis may be altered, possibly leading to changes in the flow profiles of the central vessels. The presence of smooth muscle arranged at the circumference of the foramen and in the walls of the anastomosis was demonstrated. The cardiovascular structures were innervated by nerves containing co-existing tyrosine hydroxylase, NPY and somatostatin immunoreactivities, which also occurred in neurons of the sympathetic ganglia. CGRP and substance P immunoreactive material co-existed in cardiovascular nerves, and in the nodose ganglion. In addition, bombesin, VIP and galanin immunoreactive nerves were found. Effects of neuropeptides on blood flows and blood pressures were studied in vivo. Substance P increased all blood flows measured, NPY increased the flow through the arterial anastomosis while neurotensin caused an initial decrease in the flow through the arterial anastomosis. In conclusion, there is a rich innervation of the heart and major vessels of the estuarine crocodile, including the foramen of Panizza and the arterial anastomosis. These nerves possibly regulate the distribution of blood in the cardiovascular system, which is further suggested by the results of the injection of neuropeptides.

Pre-and post-branchial blood respiratory status during acute hypercapnia or hypoxia in rainbow trout,Oncorhynchus mykiss

Simultaneous venous (pre-branchial) and arterial (post-branchial) extracorporeal blood circulations were utilized to monitor continuously the rapid and progressive effects of acute environmental hypercapnia (water partial pressure of CO2 4.8±0.2 torr) or hypoxia (water partial pressure of O2 25±2 torr) on oxygen and carbon dioxide tensions and pH in the blood of rainbow trout (Oncorhynchus mykiss). During hypercapnia, the CO2 tension in the arterial blood increased from 1.7±0.1 to 6.2±0.2 torr within 20 min and this was associated with a decrease of arterial extracellular pH from 7.95±0.03 to 7.38±0.03; the acid-base status of the mixed venous blood changed in a similar fashion. The decrease in blood pH in vivo was greater than in blood equilibrated in vitro with a similar CO2 tension indicating a significant metabolic component to the acidosis in vivo. Under normocapnic conditions, venous blood CO2 tension was slightly higher than arterial blood CO2 tension difference was abolished or reversed during the initial 25 min of hypercapnia indicating that CO2 was absorbed from the water during this period. Arterial O2 tension remained constant during hypercapnia; however, venous blood O2 tension decreased significantly (from 22.0±2.6 to 9.0±1.0 torr) during the initial 10 min. Hypercapnia elicited the release of catecholamines (adrenaline and noradrenaline) into the blood. The adrenaline concentration increased from 6±3 to 418±141 nmol · l-1 within 25 min; noradrenaline concentration increased from 3±0.5 to 50±21 nmol · l-1 within 15 min. During hypoxia arterial blood O2 tension declined progressively from 108.4±9.9 to 12.8±1.7 torr within 30 min. Venous blood O2 tension initially was stable but then decreased abruptly as catecholamines were released into the circulation. The release of catecholamines occurred concomitantly with a sudden metabolic acidosis in both blood compartments and a rise in CO2 tension in the mixed venous blood only.

Development of Adrenergic and Cholinergic Cardiac Control in Larvae of the African Clawed Frog Xenopus laevis

Cardiac responses (heart rate, stroke volume, and cardiac output) to cholinergic and adrenergic receptor stimulation were investigated in developing larvae of Xenopus laevis from Nieuwkoop and Faber (NF) stage 33/34 (newly hatched) to NF stage 53 (22 d after hatching). Effects on heart rate (fH), stroke volume (SV), and cardiac output (CO) were analyzed using in situ preparations and video‐microscopic techniques to record the continually beating heart. The results show that administration of acetylcholine to the heart decreases heart rate as early as NF stage 40. A significant reduction in SV and CO following acetylcholine administration to the heart was found at NF stages 45–53. Epinephrine had no significant effect on fH, SV, or CO at any of the stages investigated. However, an adrenergic tonus on the heart is present already at NF stage 40 (11%). This tonus increases up to a maximum (44%) at NF stages 45–47, when the maximal heart rate is found during development of X. laevis. We conclude that acetylcholine has a negative chronotropic and possibly also inotropic effect on the heart very early in development of X. laevis. We also hypothesize that the high adrenergic tonus found at NF stages 45–47 is responsible, at least in part, for the peak in heart rate seen at these stages.

Occurrence of neurotrophin receptors and transmitters in the developing Xenopus gut

Abstract. The ontogeny of gut innervation in the anuran amphibian Xenopus laevis was studied using immunohistochemistry on sections of whole larvae from NF stages 38–52. Immunoreactivity to acetylated tubulin confirmed the presence of nerve fibres as early as stages 38–39. Actin immunoreactivity was found at stage 41, indicating the presence of smooth muscle cells. Trk-like neurotrophin receptors were occasionally found in nerve fibres as soon as stages 38–39. Vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP) immunoreactivities coexisted in nerves innervating the gut wall from stages 40–41, and nitric oxide synthase (NOS) from stage 42. Substance P/neurokinin A (SP/NKA) occurred at stage 42. In all these cases, the first fibres were observed in the oesophagus. Calcitonin gene-related peptide (CGRP) was first observed in nerves at stage 48. In general, VIP/PACAP and NOS innervation was denser than the tachykinin innervation. In conclusion, the development of nerve fibres in the Xenopus gut is probably dependent on neurotrophins that may act via Trk-like receptors and occur before the gut wall is fully organised morphologically. Feeding in Xenopus larvae starts at NF stage 45. The study demonstrates that several of the transmitters investigated are expressed in the gut innervation (and in endocrine cells) prior to this stage.