It takes time to heal a broken heart: ventricular plasticity improves heart performance after myocardial infarction in rainbow trout, Oncorhynchus mykiss.

Coronary arteriosclerosis is a common feature of both wild and farmed salmonid fishes and may be linked to stress-induced cardiac pathologies. Yet, the plasticity and capacity for long-term myocardial restructuring and recovery following a restriction in coronary blood supply are unknown. Here, we analyzed the consequences of acute (3 days) and chronic (from 33 to 62 days) coronary occlusion (i.e. coronary artery ligation) on cardiac morphological characteristics and in vivo function in juvenile rainbow trout, Oncorhynchus mykiss. Acute coronary artery occlusion resulted in elevated resting heart rate and decreased inter-beat variability, which are both markers of autonomic dysfunction following acute myocardial ischemia, along with severely reduced heart rate scope (maximum-resting heart rate) relative to sham-operated trout. We also observed a loss of myocardial interstitial collagen and compact myocardium. Following long-term coronary artery ligation, resting heart rate and heart rate scope normalized relative to sham-operated trout. Moreover, a distinct fibrous collagen layer separating the compact myocardium into two layers had formed. This may contribute to maintain ventricular integrity across the cardiac cycle or, alternatively, demark a region of the compact myocardium that continues to receive oxygen from the luminal venous blood. Taken together, we demonstrate that rainbow trout may cope with the aversive effects caused by coronary artery obstruction through plastic ventricular remodeling, which, at least in part, restores cardiac performance and myocardium oxygenation.

Exposure to seawater increases intestinal motility in euryhaline rainbow trout (Oncorhynchus mykiss).

Upon exposure to seawater, euryhaline teleosts need to imbibe and desalinate seawater to allow for intestinal ion and water absorption, as this is essential for maintaining osmotic homeostasis. Despite the potential benefits of increased mixing and transport of imbibed water for increasing the efficiency of absorptive processes, the effect of water salinity on intestinal motility in teleosts remains unexplored. By qualitatively and quantitatively describing in vivo intestinal motility of euryhaline rainbow trout (Oncorhynchus mykiss), this study demonstrates that, in freshwater, the most common motility pattern consisted of clusters of rhythmic, posteriorly propagating contractions that lasted ∼1-2 min followed by a period of quiescence lasting ∼4-5 min. This pattern closely resembles mammalian migrating motor complexes (MMCs). Following a transition to seawater, imbibed seawater resulted in a significant distension of the intestine and the frequency of MMCs increased twofold to threefold with a concomitant reduction in the periods of quiescence. The increased frequency of MMCs was also accompanied by ripple-type contractions occurring every 12-60 s. These findings demonstrate that intestinal contractile activity of euryhaline teleosts is dramatically increased upon exposure to seawater, which is likely part of the overall response for maintaining osmotic homeostasis as increased drinking and mechanical perturbation of fluids is necessary to optimise intestinal ion and water absorption. Finally, the temporal response of intestinal motility in rainbow trout transitioning from freshwater to seawater coincides with previously documented physiological modifications associated with osmoregulation and may provide further insight into the underlying reasons shaping the migration patterns of salmonids.

Increased mitochondrial coupling and anaerobic capacity minimizes aerobic costs of trout in the sea.

Anadromy is a distinctive life-history strategy in fishes that has evolved independently many times. In an evolutionary context, the benefits of anadromy for a species or population must outweigh the costs and risks associated with the habitat switch. The migration of fish across the freshwater-ocean boundary coincides with potentially energetically costly osmoregulatory modifications occurring at numerous levels of biological organization. By integrating whole animal and sub-cellular metabolic measurements, this study presents significant findings demonstrating how an anadromous salmonid (i.e. rainbow trout, Oncorhynchus mykiss) is able to transform from a hyper- to hypo-osmoregulatory state without incurring significant increases in whole animal oxygen consumption rate. Instead, underlying metabolic mechanisms that fuel the osmoregulatory machinery at the organ level (i.e. intestine) are modulated, as mitochondrial coupling and anaerobic metabolism are increased to satisfy the elevated energetic demands. This may have positive implications for the relative fitness of the migrating individual, as aerobic capacity may be maintained for locomotion (i.e. foraging and predator avoidance) and growth. Furthermore, the ability to modulate mitochondrial metabolism in order to maintain osmotic balance suggests that mitochondria of anadromous fish may have been a key target for natural selection, driving species adaptations to different aquatic environments.

The presence and role of interstitial cells of Cajal in the proximal intestine of shorthorn sculpin (Myoxocephalus scorpius).

Rhythmic contractions of the mammalian gastrointestinal tract can occur in the absence of neuronal or hormonal stimulation owing to the generation of spontaneous electrical activity by interstitial cells of Cajal (ICC) that are electrically coupled to smooth muscle cells. The myogenically driven component of gastrointestinal motility patterns in fish probably also involves ICC; however, little is known of their presence, distribution and function in any fish species. In the present study, we combined immunohistochemistry and in vivo recordings of intestinal motility to investigate the involvement of ICC in the motility of the proximal intestine in adult shorthorn sculpin (Myoxocephalus scorpius). Antibodies against anoctamin 1 (Ano1, a Ca2+-activated Cl- channel), revealed a dense network of multipolar, repeatedly branching cells in the myenteric region of the proximal intestine, similar in many regards to the mammalian ICC-MY network. The addition of benzbromarone, a potent blocker of Ano1, altered the motility patterns seen in vivo after neural blockade with TTX. The results indicate that ICC are integral for the generation and propagation of the majority of rhythmic contractile patterns in fish, although their frequency and amplitude can be modulated via neural activity.

Cardiorespiratory upregulation during seawater acclimation in rainbow trout: effects on gastrointestinal perfusion and postprandial responses.

Increased gastrointestinal blood flow is essential for euryhaline fishes to maintain osmotic homeostasis during the initial phase of a transition from freshwater to seawater. However, the cardiorespiratory responses and hemodynamic changes required for a successful long-term transition to seawater remain largely unknown. In the present study, we simultaneously measured oxygen consumption rate (MO2), cardiac output (CO), heart rate (HR), and gastrointestinal blood flow (GBF) in rainbow trout (Oncorhynchus mykiss) acclimated to either freshwater or seawater for at least 6 wk. Seawater-acclimated trout displayed significantly elevated MO2 (day: 18%, night: 19%), CO (day: 22%, night: 48%), and GBF (day: 96%, night: 147%), demonstrating that an overall cardiorespiratory upregulation occurs during seawater acclimation. The elevated GBF was achieved via a combination of increased CO, mediated through elevated stroke volume (SV), and a redistribution of blood flow to the gastrointestinal tract. Interestingly, virtually all of the increase in CO of seawater-acclimated trout was directed to the gastrointestinal tract. Although unfed seawater-acclimated trout displayed substantially elevated cardiorespiratory activity, the ingestion of a meal resulted in a similar specific dynamic action (SDA) and postprandial GBF response as in freshwater-acclimated fish. This indicates that the capacity for the transportation of absorbed nutrients, gastrointestinal tissue oxygen delivery, and acid-base regulation is maintained during digestion in seawater. The novel findings presented in this study clearly demonstrate that euryhaline fish upregulate cardiovascular function when in seawater, while retaining sufficient capacity for the metabolic and cardiovascular changes associated with the postprandial response.

Tyrosine hydroxylase immunoreactivity is common in the enteric nervous system in teleosts.

Tyrosine hydroxylase (TH) is the rate-limiting enzyme in the synthesis of catecholamines and TH immunoreactivity is indicative of cells synthesising either adrenaline/noradrenaline or dopamine. In this study, the distribution of TH immunoreactivity was examined in two distantly related teleost species, zebrafish (Danio rerio) and shorthorn sculpin (Myoxocephalus scorpius). In both species, TH-immunoreactive nerve cell bodies and varicose nerve fibres were common in the myenteric plexus of the intestine. However, no TH-immunoreactive nerve cell bodies were seen in the sculpin stomach. The TH-immunoreactive nerve cell bodies seemed to constitute a larger proportion of the total enteric population in shorthorn sculpin (50 ± 5 %, n = 3067 cells) compared with zebrafish (14 ± 2 %, n = 10,163 cells). In contrast, in sculpin, the TH-immunoreactive cells were smaller than the average enteric nerve cell bodies, whereas in zebrafish, the relationship was the opposite. In developing zebrafish larvae, TH-immunoreactive nerve cell bodies were common (approx. 75 % of the total population) at 3 days post-fertilization (dpf), but decreased in numbers between 3 and 7 dpf. In conclusion, in contrast to previous studies, TH-immunoreactive intrinsic neurons are common in the fish gut. Their role and function need to be further characterized in order to understand the potential importance of this enteric subpopulation in controlling various gut functions.

Increased gastrointestinal blood flow: An essential circulatory modification for euryhaline rainbow trout (Oncorhynchus mykiss) migrating to sea.

The large-scale migrations of anadromous fish species from freshwater to seawater have long been considered particularly enigmatic, as this life history necessitates potentially energetically costly changes in behaviour and physiology. A significant knowledge gap concerns the integral role of cardiovascular responses, which directly link many of the well-documented adaptations (i.e. through oxygen delivery, water and ion transport) allowing fish to maintain osmotic homeostasis in the sea. Using long-term recordings of cardiorespiratory variables and a novel method for examining drinking dynamics, we show that euryhaline rainbow trout (Oncorhynchus mykiss) initiate drinking long before the surrounding environment reaches full seawater salinity (30-33 ppt), suggesting the presence of an external osmo-sensing mechanism. Onset of drinking was followed by a delayed, yet substantial increase in gastrointestinal blood flow through increased pulse volume exclusively, as heart rate remained unchanged. While seawater entry did not affect whole animal energy expenditure, enhanced gastrointestinal perfusion represents a mechanism crucial for ion and water absorption, as well as possibly increasing local gastrointestinal oxygen supply. Collectively, these modifications are essential for anadromous fish to maintain homeostasis at sea, whilst conserving cardiac and metabolic scope for activities directly contributing to fitness and reproductive success.

Effects of feeding on in vivo motility patterns in the proximal intestine of shorthorn sculpin (Myoxocephalus scorpius).

This is the first study to catalogue the diverse array of in vivo motility patterns in a teleost fish and how they are affected by feeding. Video recordings of exteriorised proximal intestine from fasted and fed shorthorn sculpin (Myoxocephalus scorpius) were used to generate spatio-temporal maps to portray and quantify motility patterns. Propagating and non-propagating contractions were observed to occur at different frequencies and durations. The most apparent difference between the feeding states was that bands of relatively high amplitude contractions propagating slowly in the anal direction were observed in all fasted fish (N=10) but in only 35% of fed fish (N=11). Additionally, fed fish displayed a reduced frequency (0.21±0.03 versus 0.32±0.06 contractions min(-1)) and rhythmicity of these contractions compared with fasted fish. Although the underlying mechanisms of these slow anally propagating contractions differ from those of mammalian migrating motor complexes, we believe that they may play a similar role in shorthorn sculpin during the interdigestive period, to potentially remove food remnants and prevent the establishment of pathogens. 'Ripples' were the most prevalent contraction type in shorthorn sculpin and may be important during mixing and absorption. The persistence of shallow ripples and pendular movements of longitudinal muscle after tetrodotoxin (1 µmol l(-1)) treatment suggests these contractions were myogenic in origin. The present study highlights both similarities and differences in motility patterns between shorthorn sculpin and other vertebrates, as well as providing a platform to examine other aspects of gastrointestinal functions in fish, including the impact of environmental changes.

Aerobic scope fails to explain the detrimental effects on growth resulting from warming and elevated CO2 in Atlantic halibut.

As a consequence of increasing atmospheric CO2, the world's oceans are becoming warmer and more acidic. Whilst the ecological effects of these changes are poorly understood, it has been suggested that fish performance including growth will be reduced mainly as a result of limitations in oxygen transport capacity. Contrary to the predictions given by the oxygen- and capacity-limited thermal tolerance hypothesis, we show that aerobic scope and cardiac performance of Atlantic halibut (Hippoglossus hippoglossus) increase following 14-16 weeks exposure to elevated temperatures and even more so in combination with CO2-acidified seawater. However, the increase does not translate into improved growth, demonstrating that oxygen uptake is not the limiting factor for growth performance at high temperatures. Instead, long-term exposure to CO2-acidified seawater reduces growth at temperatures that are frequently encountered by this species in nature, indicating that elevated atmospheric CO2 levels may have serious implications on fish populations in the future.

Metabolic scope and interspecific competition in sculpins of Greenland are influenced by increased temperatures due to climate change.

Ongoing climate change has led to an increase in sea surface temperatures of 2-4°C on the west coast of Greenland. Since fish are ectothermic, metabolic rate increases with ambient temperature. This makes these animals particularly sensitive to changes in temperature; subsequently any change may influence their metabolic scope, i.e. the physiological capacity to undertake aerobically challenging activities. Any temperature increase may thus disrupt species-specific temperature adaptations, at both the molecular level as well as in behavior, and concomitant species differences in the temperature sensitivity may shift the competitive balance among coexisting species. We investigated the influence of temperature on metabolic scope and competitive ability in three species of marine sculpin that coexist in Greenland coastal waters. Since these species have different distribution ranges, we hypothesized that there should be a difference in their physiological response to temperature; hence we compared their metabolic scope at three temperatures (4, 9 and 14°C). Their competitive ability at the ambient temperature of 9°C was also tested in an attempt to link physiological capacity with behaviour. The Arctic staghorn sculpin, the species with the northernmost distribution range, had a lower metabolic scope in the higher temperature range compared to the other two species, which had similar metabolic scope at the three temperatures. The Arctic staghorn sculpin also had reduced competitive ability at 9°C and may thus already be negatively affected by the current ocean warming. Our results suggest that climate change can have effects on fish physiology and interspecific competition, which may alter the species composition of the Arctic fish fauna.

Calbindin immunoreactivity in the enteric nervous system of larval and adult zebrafish (Danio rerio).

Calbindin is a calcium-binding protein, commonly found in certain subpopulations of the enteric nervous system in mammals. Recently, calbindin-immunoreactive enteric neurons have also been demonstrated in shorthorn sculpin (Myoxocephalus scorpius). In the present study, calbindin immunoreactivity has been investigated in the gut of adult and larval zebrafish (Danio rerio) and differences and similarities between the two species are discussed. Calbindin immunoreactivity is present in 40%-50% of all enteric neurons in adult zebrafish. It first appears at 3 days post-fertilisation (dpf) and is present in all regions of the gut by 13 dpf. Calbindin-immunoreactive nerve cell bodies do not differ in size from calbindin-negative cells. Zebrafish calbindin-immunoreactive neurons are serotonin-negative, with at least some being choline acetyltransferase (ChAT)-positive, in contrast to the sculpin in which cells are generally smaller than the average enteric neuron and are serotonin-positive and ChAT-negative. These findings further emphasise the importance of comparative studies for understanding the diversity of chemical coding in the enteric nervous system of fish and other vertebrates. Improved knowledge of the role of the enteric nervous system is also essential for future studies of gut activity with regard to zebrafish being used as a model organism.

Calbindin-immunoreactive cells in the fish enteric nervous system.

Calbindin is present in a large proportion of the intrinsic primary afferent neurons (IPANs) in the mammalian gut. Little is known about either calbindin or IPANs in fish. In the present study, calbindin immunoreactivity was investigated in the enteric nervous system of the teleost shorthorn sculpin (Myoxocephalus scorpius). Calbindin-immunoreactive nerve cell bodies and nerve fibres were present in all the gut regions except the cardiac stomach. The highest proportion was found in the proximal intestine where calbindin-immunoreactive cells constituted 59±6% (N=3) of the total Hu C/D-immunoreactive myenteric nerve cell population. In other regions, calbindin-immunoreactive cells constituted around 30% of the total population. The cells were generally multipolar with one long axon. The size distribution differed significantly between calbindin-positive and calbindin-negative cells in each of the three animals examined. Calbindin-positive neurons in the proximal intestine had a mean cross-sectional soma area of 163±73µm(2) (n=183 cells) while calbindin-negative cells were 348±221µm(2) (n=127 cells). Calbindin immunoreactivity colocalised to a large extent with serotonin immunoreactivity, but not with choline acetyltransferase (ChAT)-immunoreactivity. Thus, the calbindin-immunoreactive nerve cell population in the shorthorn sculpin gut seems to constitute a homogenous subpopulation of the enteric neurons, at least when considering the size and content of some transmitters. Whether markers other than serotonin and ChAT would differentiate the population remains to be tested. In conclusion, the calbindin-immunoreactive cells in the sculpin differ from mammalian IPANs with regard to several parameters and future functional studies could hopefully add information about the role of this large group of cells in the fish enteric nervous system.

Autonomic control of gut motility: a comparative view.

Gut motility is regulated to optimize food transport and processing. The autonomic innervation of the gut generally includes extrinsic cranial and spinal autonomic nerves. It also comprises the nerves contained entirely within the gut wall, i.e. the enteric nervous system. The extrinsic and enteric nervous control follows a similar pattern throughout the vertebrate groups. However, differences are common and may occur between groups and families as well as between closely related species. In this review, we give an overview of the distribution and effects of common neurotransmitters in the vertebrate gut. While the focus is on birds, reptiles, amphibians and fish, mammalian data are included to form the background for comparisons. While some transmitters, like acetylcholine and nitric oxide, show similar distribution patterns and effects in most species investigated, the role of others is more varying. The significance for these differences is not yet fully understood, emphasizing the need for continued comparative studies of autonomic control.

Autonomic control of glands and secretion: a comparative view.

The autonomic nervous system together with circulating and local hormones control secretion from glands. This article summarizes histochemical and functional studies on the autonomic innervation and control of secretory glands in non-mammalian vertebrates, including secretion of saliva in the mouth and gastric acid in the stomach, secretion of enzymes and bicarbonate from the pancreas and gut wall, secretion of mucus in the gut epithelium and onto the skin, and salt secretion from salt glands and rectal glands. Cholinergic and adrenergic nerves, directly or indirectly, in combination with different types of peptidergic and other nerves appear to innervate gland tissues and affect secretion in all investigated species.

Glial cells revealed by GFAP immunoreactivity in fish gut.

Glial fibrillary acidic protein (GFAP) is a commonly used marker to identify enteric glia in the mammalian gut. Little is however known about enteric glia in other vertebrates. The aim of the present study was to examine the distribution of GFAP immunoreactivity in adult and developing fish. In adult shorthorn sculpin (Myoxocephalus scorpius) and zebrafish (Danio rerio), GFAP immunoreactivity was seen in the myenteric plexus in all regions of the gut. Co-staining for the neuronal markers Hu C/D and acetylated tubulin showed that GFAP immunoreactivity was not associated with nerves. GFAP immunoreactivity was predominantly seen in processes with few glial cell bodies being demonstrated in adult fish. GFAP immunoreactivity was also found in the gut in larval zebrafish from 3 days post-fertilisation, i.e. at approximately the same time that differentiated enteric nerve cells first occur. Immunoreactivity was most prominent in areas with no or a low density of Hu-immunoreactive nerve cell bodies, indicating that the developing glia follows a different pattern from that of enteric neurons. The results suggest that GFAP can be used as a marker for enteric glia in fish, as in birds and mammals. The distribution of GFAP immunoreactivity implies that enteric glia are widespread in the fish gastrointestinal tract. Glia and neurons diverge early during development of the gastrointestinal tract.

Nervous and humoral catecholaminergic control of blood pressure and cardiac performance in the Antarctic fish Pagothenia borchgrevinki.

The role of circulating and neural catecholamines for cardiovascular control in the Antarctic fish Pagothenia borchgrevinki was studied in vivo using pharmacological tools and with immunohistochemistry on isolated tissues. Adrenergic nerve blockade with bretylium decreased dorsal aortic pressure (P(da)) and systemic vascular resistance (R(sys)), while cardiac output (Q) did not change. The blockade of alpha-adrenoceptors with phentolamine reduced P(da) and R(sys) further, revealing that vasomotor tone was influenced by circulating catecholamines in bretylium treated fish. The physiological evidence for an adrenergic nervous control of the vasculature was corroborated by the presence of tyrosine hydroxylase (TH)-immunoreactive fibres associated with blood vessels in spleen, gonads and gastrointestinal tract. TH-immunoreactive fibres were not observed in the atrium and ventricle, but a dense population of TH-immunoreactive fibres was apparent in the bulbus arteriosus. The present study suggests that an adrenergic nervous mechanism is responsible for maintaining vasomotor tone in P. borchgrevinki. While experiments failed to demonstrate a tonic adrenergic nervous influence affecting cardiac performance, an adrenergic nervous control of bulbar compliance may be essential for optimizing gill blood flow dynamics in this species, which has a high relative stroke volume and displays profound changes in stroke volume in vivo.

Postprandial changes in enteric electrical activity and gut blood flow in rainbow trout (Oncorhynchus mykiss) acclimated to different temperatures.

Enteric electrical activity, cardiac output and gut blood flow were measured in rainbow trout (Oncorhynchus mykiss) acclimated to either 10 degrees C or 16 degrees C. Enteric electrical activity showed, in both the fasted and postprandial state, a distinct pattern with clusters of burst-like events interspersed by silent periods. The frequency of electrical events increased postprandially for both acclimation groups. Event frequency increased from 3.0+/-0.5 to 9.6+/-1.4 events min(-1) and from 5.9+/-0.9 to 11.8+/-2.0 events min(-1) in the 10 degrees C and 16 degrees C groups, respectively. Similarly, the number of events per cluster increased postprandially for both acclimation groups. Gut blood flow, cardiac output and heart rate increased after feeding. The gut blood flow significantly increased in both groups and peaked at 257+/-19% and 236+/-22% in the 10 degrees C and 16 degrees C groups, respectively. There was a strong correlation between the number of events and gut blood flow at both temperatures. Comparison between the two groups showed that fish acclimated to 16 degrees C may have an increased cost of sustaining the basal activity of the gut compared with the group acclimated to 10 degrees C. In conclusion, we have for the first time measured enteric electrical activity in vivo in a fish species and we have also demonstrated a strong correlation between gut blood flow and enteric electrical activity in fasted and postprandial fish.

Autonomic innervation of the fish gut.

The enteric nervous system follows a similar overall arrangement in all vertebrate groups. In fish, the majority of nerve cell bodies are found in the myenteric plexus, innervating muscles, blood vessels and glands. In this review, I describe similarities and differences in size, shape and transmitter content in enteric neurons in different fish species and also in comparison with other vertebrates, foremost mammals. The use of different histological and immunochemical methods is reviewed in a historical perspective including advantages and disadvantages of different methods. Lately, zebrafish have become an important model species for developmental studies of the nervous system, including the enteric nervous system, and this is briefly discussed. Finally, examples of how the enteric nervous system controls gut activity in fish is presented, focussing on the effect on gastrointestinal motility.