Lynne U. Sneddon

Do fishes have nociceptors? Evidence for the evolution of a vertebrate sensory system.

Nociception is the detection of a noxious tissue–damaging stimulus and is sometimes accompanied by a reflex response such as withdrawal. Pain perception, as distinct from nociception, has been demonstrated in birds and mammals but has not been systematically studied in lower vertebrates. We assessed whether a fish possessed cutaneous nociceptors capable of detecting noxious stimuli and whether its behaviour was sufficiently adversely affected by the administration of a noxious stimulus. Electrophysiological recordings from trigeminal nerves identified polymodal nociceptors on the head of the trout with physiological properties similar to those described in higher vertebrates. These receptors responded to mechanical pressure, temperatures in the noxious range (more than 40°C) and 1% acetic acid, a noxious substance. In higher vertebrates nociceptive nerves are either A–delta or C fibres with C fibres being the predominating fibre type. However, in the rainbow trout A–delta fibres were most common, and this offers insights into the evolution of nociceptive systems. Administration of noxious substances to the lips of the trout affected both the physiology and the behaviour of the animal and resulted in a significant increase in opercular beat rate and the time taken to resume feeding, as well as anomalous behaviours. This study provides significant evidence of nociception in teleost fishes and furthermore demonstrates that behaviour and physiology are affected over a prolonged period of time, suggesting discomfort.

The evidence for pain in fish: the use of morphine as an analgesic

Abstract This paper discusses the evidence for pain perception in fish and presents new data on morphine analgesia in fish. Recent anatomical and electrophysiological studies have demonstrated that fish are capable of nociception, the simple detection of a noxious, potentially painful stimulus and the reflex response to this. To prove pain perception, it must be demonstrated that an animals behaviour is adversely affected by a potentially painful event and this must not be a reflex response. The present study examined the acute effects of administering a noxious chemical to the lips of rainbow trout ( Oncorhynchus mykiss ) to assess what changes occurred in behaviour and physiology. There was no difference in swimming activity or use of cover when comparing the noxiously stimulated individuals with the controls. The noxiously treated individuals performed anomalous behaviours where they rocked on either pectoral fin from side to side and they also rubbed their lips into the gravel and against the sides of the tank. Opercular beat rate (gill or ventilation rate) increased almost double fold after the noxious treatment whereas the controls only showed a 30% increase. Administering morphine significantly reduced the pain-related behaviours and opercular beat rate and thus morphine appears to act as an analgesic in the rainbow trout. It is concluded that these pain-related behaviours are not simple reflexes and therefore there is the potential for pain perception in fish.

Plasticity in animal personality traits: does prior experience alter the degree of boldness?

Theoreticians predict that animal ‘personality’ traits may be maladaptive if fixed throughout different contexts, so the present study aimed to test whether these traits are fixed or plastic. Rainbow trout (Onchorhyncus mykiss) were given emboldening or negative experiences in the forms of watching bold or shy individuals responding to novelty or winning or losing fights to examine whether prior experience affected boldness. Bold individuals that lost fights or watched shy demonstrators became more shy by increasing their latency to approach a novel object, whereas shy observers that watched bold demonstrators remained cautious and did not modify their responses to novelty. Shy winners became bolder and decreased their latency to approach a novel object, but shy losers also displayed this shift. In comparison, control groups showed no change in behaviour. Bold fishes given negative experiences reduced their boldness which may be an adaptive response; however, shy fishes may base their strategic decisions upon self-assessment of their relative competitive ability and increase their boldness in situations where getting to resources more quickly ensures they outcompete better competitors.

Weapon size versus body size as a predictor of winning in fights between shore crabs, Carcinus maenas (L.)

Relative body size (carapace width) and weapon size (chela length) were used as indicators of resource holding potential (RHP) in the agonistic behaviour of male shore crabs, Carcinus maenas (L.). Weapon size was found to be a more reliable predictor of the outcome of pairwise fights than body size. Crabs with longer chelae than their opponents were more likely to win fights than crabs with relatively larger bodies. Body size had less influence on the outcome of fights. Relative body and weapon size did not influence initiation of contests but did affect the likelihood of winning; however, this was significant only for weapon size. Winning crabs had heavier claws with greater surface area than losing crabs. There was no relationship between relative size and fight duration. The frequency of cheliped display increased with chela length and win- ners performed significantly more displays than losers.

Novel object test: examining nociception and fear in the rainbow trout.

This study aimed to assess fear responses to a novel object while experiencing a noxious event to determine whether nociception or fear will dominate attention in a fish in novel object testing paradigm. This experimentally tractable animal model was used to investigate (1) the degree of neophobia to a novel object while experiencing noxious stimulation, (2) the response of the fish after removing the fear-causing event by using a familiar object, and (3) the effects of removing the nociceptive response by morphine administration and examining the response to a novel object. Control animals displayed a classic fear response to the novel objects and spent most of their time moving away from this stimulus, as well as showing an increase in respiration rate when the novel object was presented. In contrast, noxiously stimulated animals spent most of their time in close proximity to the novel object and showed no additional increase in respiration rate to novel object presentation. There was evidence of a slight hypoalgesia in noxiously stimulated animals. The responses to familiar objects demonstrated that by familiarizing the animal with the object, fear was removed from the experiment. Both control and noxiously treated animals responded in similar ways to a novel object by spending the majority of their time in close proximity. Treatment with morphine reduced effects of noxious stimulation and appears to be an effective analgesic. After morphine administration, the acid-injected animals showed a neophobic response to a novel object and this was similar to the response of the control fish, with a similar amount of time spent moving away from the object and an increase in ventilation in response to the novel object. Morphine affected the fear response because both groups approached the novel object more quickly than the non-morphine controls. These results suggest that nociception captures the animals attention with only a relatively small amount of attention directed at responding to the fear of the novel object.

Trigeminal Somatosensory Innervation of the Head of a Teleost Fish with Particular Reference to Nociception

Trigeminal somatosensory receptors have not been characterised in teleost fish and studies in elasmobranchs have failed to identify nociceptors. The present study examined the trigeminal nerve of a teleost fish, the rainbow trout (Oncorhynchus mykiss) to determine what types of somatosensory receptors were present on the head of the trout specifically searching for nociceptors. Single unit recordings were made from receptive fields on the head of the fish innervated by the trigeminal nerve. Each receptive field was tested for sensitivity to mechanical, thermal and chemical stimulation. Five different receptor types were found: fast adapting receptors responding to mechanical stimulation; slowly adapting receptors responding to mechanical stimuli; polymodal nociceptors responding to mechanical, noxious thermal and chemical stimulation; mechanothermal nociceptors responding to mechanical stimulation and noxious heat; and mechanochemical receptors responsive to mechanical and chemical stimulation. Mechanical thresholds, receptive field diameter, conduction velocities and thermal thresholds of the receptors were determined and there was no significant difference between the receptor types in terms of these properties. Three shapes of action potential (AP) were recorded from these receptors: type 1 with no inflexion; type 2 with an inflexion on depolarisation; and type 3 with an inflexion on repolarisation. Conduction velocity, amplitude and duration of the APs, afterhypolarisation amplitude and duration, as well as the maximum rate of depolarisation were measured for each action potential type. No major differences were found when making comparisons within receptor type and between receptor types. The fish nociceptors had similar physiological properties to nociceptors found in higher vertebrates.

Anatomical and electrophysiological analysis of the trigeminal nerve in a teleost fish, Oncorhynchus mykiss.

The trigeminal nerve in the rainbow trout, Oncorhynchus mykiss, was examined for the presence of A-delta and C fibres. Sections of the three branches of the trigeminal nerve were found to comprise a range of fibre types including A-delta and C fibres. The size range of the cell bodies of the trigeminal ganglion reflected the fibre range since they correlated with the size range of axons in the nerve branches. Electrophysiological recordings of evoked activity from the ganglion confirmed the presence of these fibre types and the proportion of these mirrored the proportion of fibre types in the anatomical analyses. A-beta fibres were most common followed by A-delta fibres, then A-alpha fibres with C fibres being the fewest fibre type found. In higher vertebrates, A-delta and C fibres in the trigeminal nerve convey both somatosensory and nociceptive information to the brain. The evolutionary significance of these results is discussed as well as the potential for nociceptive capability in a lower vertebrate.

Defining and assessing animal pain

The detection and assessment of pain in animals is crucial to improving their welfare in a variety of contexts in which humans are ethically or legally bound to do so. Thus clear standards to judge whether pain is likely to occur in any animal species is vital to inform whether to alleviate pain or to drive the refinement of procedures to reduce invasiveness, thereby minimizing pain. We define two key concepts that can be used to evaluate the potential for pain in both invertebrate and vertebrate taxa. First, responses to noxious, potentially painful events should affect neurobiology, physiology and behaviour in a different manner to innocuous stimuli and subsequent behaviour should be modified including avoidance learning and protective responses. Second, animals should show a change in motivational state after experiencing a painful event such that future behavioural decision making is altered and can be measured as a change in conditioned place preference, self-administration of analgesia, paying a cost to access analgesia or avoidance of painful stimuli and reduced performance in concurrent events. The extent to which vertebrate and selected invertebrate groups fulfil these criteria is discussed in light of the empirical evidence and where there are gaps in our knowledge we propose future studies are vital to improve our assessment of pain. This review highlights arguments regarding animal pain and defines criteria that demonstrate, beyond a reasonable doubt, whether animals of a given species experience pain.

Metabolic consequences of agonistic behaviour: crab fights in declining oxygen tensions.

The energetic consequences of fighting, which may depend on environmental conditions, can be an important factor shaping contest strategy and duration. Energy expenditure may be costly to fitness because it depletes reserves that could otherwise have been allocated to reproduction, and metabolites are produced that may constrain subsequent activities. We examined the variation in the metabolic consequences of fighting in relation to hypoxia. Contests were staged between pairs of size-matched male shore crabs Carcinus maenas L., under a range of water oxygen tensions (between 10 and 100% oxygen saturation) which crabs experience in their natural habitat. Fighting under normoxic and hypoxic conditions resulted in significantly elevated concentrations of haemolymph metabolites (L-lactate and glucose) compared with crabs at rest. However, these concentrations were much lower than in crabs that had been walking on a treadmill. Glycogen concentrations differed only under hypoxic conditions: glycogen stores were reduced in crabs after fighting and this reduction was similar to that after exercise on a treadmill. Contests were shorter when they were staged below a water P o2of 6.7 kPa ( approximately 30% normoxia). As water oxygen tensions were reduced, fighting crabs had greater concentrations of L-lactate and glucose in their blood and tissues whilst glycogen stores were reduced. Fights became shorter when crabs were exposed to severe hypoxia (P o2=2 kPa) for increasing lengths of time, and blood L-lactate concentrations increased. The results suggest that as fights progressed, crabs experienced an increasing metabolic debt, in the form of accumulation of L-lactate and a reduction in energy stores, which was amplified by hypoxic conditions. Copyright 1999 The Association for the Study of Animal Behaviour.

Nociception in fish: stimulus-response properties of receptors on the head of trout Oncorhynchus mykiss.

This study examined stimulus-response properties of somatosensory receptors on the head of rainbow trout, Oncorhynchus mykiss, using extracellular recording from single cells in the trigeminal ganglion. Of 121 receptors recorded from 39 fish, 17 were polymodal nociceptors, 22 were mechanothermal nociceptors, 18 were mechanochemical receptors, 33 were fast adapting mechanical receptors and 31 were slowly adapting mechanical receptors. Mechanical thresholds were higher in polymodal nociceptors than in either slowly adapting or fast adapting mechanical receptors, whereas thermal thresholds of mechanothermal nociceptors were higher than those of polymodal nociceptors. Polymodal nociceptors and mechanochemical receptors gave similar responses to topical applications of acid. All receptor types except mechanothermal nociceptors showed an increase in peak firing frequency with increased strength of mechanical stimulation, with evidence of response saturation at higher intensities. Mechanothermal, but not polymodal, nociceptors showed an increase in firing response to increased temperature. None out of 120 receptors tested gave any response to the temperature range +4 degrees C to -7 degrees C, indicating an absence of cold nociceptors. Attempts to evoke sensitization of receptors using chemical or heat stimuli were unsuccessful, with receptors showing either a return to control responses or irreversible damage. Comparisons are made between somatosensory receptors characterized here in a fish and those of higher vertebrates.