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Home , Animal welfare science, Live food, Pain in invertebrates

PostDoc Journal Journal of Postdoctoral Research Vol. 1, No. 2, February 2013 www.postdocjournal.com

The welfare of in research: Can science's next generation improve their lot? Robyn J. Crook Department of Integrative Biology and Pharmacology, University of Texas Health Science Center, Houston, TX 77054 Email: [email protected]

Abstract have a long history of use in scientific research but there has been little concern for their welfare until very recently. Unlike vertebrate research animals, whose uses are closely regulated, invertebrate animals are minimally protected. In some countries regulations extend to a few species, but the vast majority of invertebrate animals can be used in research with no oversight, protections or legal regulation. Whether this is cause for concern depends on the ability of invertebrate animals to experience , or distress as a result of husbandry and experimental procedures. To date there is minimal evidence that invertebrate animals are capable of experiencing such affective states, but this is largely due to very little experimental effort devoted to testing such hypotheses. In this article I review current regulatory guidelines for use of invertebrate animals and their relevance to species of varying neural and behavioural complexity. I define some simple steps that postdoctoral and other young researchers can take to improve the welfare of research animals within their own laboratories, and provide a framework for contributing more broadly to the field of welfare-based research by publishing their observations when improvements have had a positive effect on their animals and their research.

Introduction and junior technicians – form the bulk of the hands-on workers in most laboratories. It is these Almost all researchers who use experimental researchers who perform most of the day-to-day animals will encounter questions at some point in tasks involving research animals, provide training their careers regarding the ethics of to new lab members, and who will eventually research. For researchers working with become leaders of their own research programs invertebrate animals these questions can be and laboratories, within which they will preside difficult to answer; the welfare of invertebrate over the use of their own research animals. Post- animals has received comparably little study, and doctoral researchers are often the most senior there are few if any regulations on their use personnel involved day-to-day in research, and (Harvey-Clark, 2011). But this is beginning to we are therefore well placed to direct, influence, change, and within the past decade concerted and ultimately improve the way we utilize efforts have been made to extend regulatory research animals. protection to invertebrate animals (see below). It is likely that within the coming years legislative How can postdoctoral researchers and other efforts will expand both their geographic and young scientists contribute to the effort to phylogenetic reach, ultimately regulating improve welfare of invertebrates used in research on multiple species of invertebrates in scientific research? In this article I define a dual most countries in which scientific research approach that includes efforts both at the occurs. individual level to change and improve their own laboratories’ practices, and in the larger arena of Young scientists - postdocs, graduate students welfare-based research, identifying which 10 Journal of Postdoctoral Research 2013: 9-20 invertebrates have the capacity to suffer pain or loosely organised nervous systems (e.g. distress, how this can be minimized, and how cnidarians, planarians, ), there is regulations governing the use of these species minimal concern about welfare. Animals with might be best formulated. more complex sensory abilities and larger nervous systems deserve more extensive Invertebrates in scientific research consideration; complexity and size of the nervous Invertebrates have a long history of scientific use. system is probably the most appropriate measure The most numerous invertebrates in modern by which we define those invertebrate animals laboratories are certainly on which efforts to improve welfare should focus melanogaster, an , and Caenorhabditis (Mather, 2011). elegans, a , but many other species contained in most invertebrate phyla are The taxa that generate the most concern represented to lesser degrees in a wide variety of therefore are the (e.g. , scientific enterprises (Wilson-Sanders, 2011). arachnids, , etc.), and molluscs (e.g., , and ), because these Along with Drosophila and C. elegans, which are invertebrates have typically the most complex used widely, developmental biologists utilize and behaviors (although there is many insects, molluscs, turnicates and considerable variations among them). My own echinoderms. Neurobiologists work extensively experimental efforts are confined within these with gastropods and cephalopods (molluscs), but two phyla, so many of the examples given in this also insects, and crustaceans article come from experience with crustaceans (arthropods), worms () and and cephalopods, but I would nonetheless urge (planarians). Immunologists utilize the self- researchers to consider welfare beyond these recognition systems of colonial invertebrates two phyla alone. such as ascidians (chordates) and corals and anemones (cnidarians) to understand The current regulatory environment fundamental mechanisms of immune functions, Welfare of research animals is not a new and study parasitism and infectious disease concern; the first act of a parliament governing processes using insects, marine shrimp the rightful uses of animals in scientific (arthropods) and roundworms (nematodes). experimentation was passed in 1876 (the UK Biomedical studies using drosophila and C. Act), and since then many elegans have yielded important insights into countries have enacted their own legislation that diseases of addiction, cancer and genetic seeks to protect animals used in research from diseases. Sensory biologists use many unreasonable harm, distress and pain. But in each invertebrate species in behavioural and case the term ‘animal’ is either implicitly or physiological studies, including crickets, , explicitly defined as a vertebrate animal (and in , flies and (arthropods), and some cases, only subsets thereof). The idea that , , squid and snails (molluscs). invertebrate animals may also deserve similar Ethologists use many species, including consideration is a rather newer concept. honeybees, ants, roaches and moths, along with Vertebrate animals used in scientific research are crustaceans and, to a lesser degree, molluscs protected by strict regulation governing their such as gastropods and cephalopods (Wilson- housing, handling, breeding and all experimental Sanders, 2011). procedures, including analgesia and euthanasia (for example, the Act, USA, Among these species (and the many not included 1966). The primary ethos underpinning such among the examples above) there is a wide range regulation is that vertebrate animals have similar of neural and behavioural complexity, and for (although not necessarily identical) capacity to those animals with very small, primitive or feel distress and pain that we have as humans,

Robyn J. Crook 11 and therefore for our research to be ethically UK law requires approval only for the use of the acceptable, we must conduct our studies in such common , Octopus vulgaris. Canada’s a way that minimizes these negative experiences CCAC expressly differentiates between (Allen, 2004; Carbone, 2004; Kaliste, 2004; ‘invertebrates’, which are placed in the tier of Baumans, 2005). lowest concern for invasive procedures, and ‘cephalopods and other higher invertebrates’ The use of invertebrate animals, by contrast, is which may be included in the higher tiers, but still very minimally regulated. Only recently have does not specify what these ‘higher there been efforts in a few countries to extend invertebrates’ might be. The EU’s new directive, some of the same protections given to vertebrate implemented in 2012, includes all animals to some invertebrates, but these cover molluscs but no other invertebrates, despite only a few species, and in some cases are originally also considering crustaceans. Although grounded on minimal knowledge of the animals’ there is no published evidence to date that . This general exclusion of cephalopod molluscs are more capable of invertebrate animals from legal protection is experiencing negative affective states than any predicated on the assumption that invertebrates, other invertebrate, or that their emotive capacity being in the majority simpler, behaviourally and is similar to that of vertebrate animals’, their neuroantomically, than vertebrates, cannot large brains and complex behavioural repertories experience pain or distress (Eisemann et al., are used as supporting evidence for this 1984; Fiorito, 1986; Kavaliers, 1988; Mather, possibility (Mather, 2001, 2011). However, the 2001; Sømme, 2005) and therefore our relatively lack of solid evidence underlying the current minimal concern for them is justified (Kellert, guidelines must be addressed in the near future if 1993). This same assumption had previously been we expect regulations to be functional in the long applied to various ‘lower’ vertebrates – , term. fish and . For each of these, careful research has provided evidence that these What is important when considering welfare of omissions were unwarranted (Stevens, 1988; invertebrates? Sneddon et al., 2003; Sneddon, 2004; Ross and Welfare considerations for invertebrates in Ross, 2008; Mosley, 2011), and legislation has research are largely similar to those for generally been amended to include them (e.g., vertebrate animals, although there is often far the US Public Health Service’s Policy on Humane less information readily available to inform those Care and Use of Laboratory Animals has been concerns. Concerns about appropriate housing, amended to include both adult and larval feeding, social interactions and handling are amphibians and fish). As we begin to consider valid, but by far the most common concern is that whether invertebrates too encompass capacity to of pain, distress and suffering during procedures. experience emotive and affective states, this same slow change will almost certainly proceed or pain? further into the invertebrate realm. Most animals having a can detect noxious or damaging stimuli, and will make Despite scarce experimental evidence currently avoidance or withdrawal motions when for affective states, necessary to enable the stimulated. This is nociception - a capacity to attachment of emotive value onto physical react to tissue damage or impending damage sensations, in any invertebrate animal (Mason, with activation of sensory and motor pathways, 2011; Mendl et al., 2011) , restrictions on the use with or without conscious awareness. Activity in of selected species are already in place in the UK nociceptive sensory pathways usually results in (Animals (Scientific Procedures) Act, 1993), reflexive behavioral responses and may or may Canada (Canadian Council on Animal Care (199) not also produce other, higher-order responses. and in the EU (Directive 2010/63/EU, 2010). The Reflexive withdrawal responses tend to be

12 Journal of Postdoctoral Research 2013: 9-20 mediated by very simple sensorimotor circuits that they occur in many others, but no optimized for speed and reliability and can occur invertebrate has been shown to experience pain. without input from higher processing centers, Because nociception does not necessarily involve although more complex escape and avoidance any negative ‘feeling’ about the sensation, and is behaviors involve more complex neural circuits by definition not aversive, alone it need not be (Walters, 1994; Chase, 2002). Even in humans the grounds for concern. However, for any animal, initial reflexive response to a noxious stimulus is including invertebrate animals, with brains even sometimes faster than can be consciously plausibly capable of attaching emotive value to perceived, and can sometimes be nociceptive experiences (i.e., can suffer and feel activated without conscious sensation pain), preventing activation of nociceptive (Adriaensen et al., 1980). Invertebrates that lack pathways during experimental procedures should appropriate higher processing centers in their be paramount. central brains may be capable of only this rapid, unconscious processing. Improving welfare of research invertebrates: A two-pronged approach for young investigators The definition of pain widely accepted by scientific investigators is “an unpleasant sensory In the remainder of this article I describe a two- and emotional experience associated with actual pronged approach for young investigators aiming or potential tissue damage, or described in terms to improve the welfare of invertebrate animals of such damage” (Merskey and Bogduk, 1994). used in research. The first involves efforts at the The emotional component required by this local level to make changes to husbandry and definition of pain makes its identification in other experimental procedures for animals in our direct species, and especially in species quite different care, and the second encompasses wider efforts from humans, extremely difficult, if not in welfare-focused research. impossible. is usually defined in terms of conscious experience (e.g., Izard, 2009), and In the laboratory while evidence of consciousness in some animals For postdocs and other young investigators, is available, proof of consciousness is not (Allen, changing an existing laboratory culture can be 2004). It is therefore important to distinguish rather difficult. However, there are a number of between nociception as detection of a noxious relatively small and easy steps a researcher can stimulus (which can be recognized scientifically take to make a substantial difference to the by unambiguous behavioral and neural and deaths of their research animals. There are responses), and pain as the unpleasant feeling two primary aspects of welfare within the associated with that stimulus (and inferred by laboratory environment – husbandry and behavioral and neural responses of uncertain experimental procedures – both of which provide relation to consciousness). Pain requires neural scope for welfare-related improvements. circuitry that incorporates additional functions, some of which might entail highly complex Husbandry processing by very large numbers of . Whether any invertebrate species might meet The first step to improving husbandry procedures these minimal criteria for experiencing the (such as housing and feeding) is to understand emotive aspect of pain is debatable, and there the biology of the species you are using. The have been only a few studies aimed at testing this natural behaviours and habits of the animals directly (Elwood and Appel, 2009; Elwood, 2011). should form the basis for husbandry regimens in the lab. Assembling a checklist of simple Nociception and nociceptive sensory neurons questions about current husbandry procedures is have been described in many invertebrate a useful means of identifying potential species (Smith and Lewin, 2009) and it is likely deficiencies, and I provide some general

Robyn J. Crook 13 suggestions here; species-specific lists should be though it might be more convenient. For developed wherever possible. example, octopuses can be highly aggressive towards conspecifics both in the wild and in Is the housing arrangement appropriate? (Bradley, 1974; Hanlon and Forsythe, Housing arrangements must balance comfort of 1985; Huffard et al., 2010). Housing multiple animals against convenience for experimenters; octopuses in a closed system is unavoidable for completely naturalistic housing is rarely labs without access to running seawater, thus appropriate or feasible for experimental animals. maintaining olfactory isolation is all but However, some considerations for naturalistic impossible. In the case where water must be habitat and abiotic features for each species can recirculated, including visual if not olfactory be incorporated into artificial housing. barriers between animals may still significantly enhance welfare and reduce stress. In octopuses If your animals are primarily nocturnal or prefer stress levels can be monitored noninvasively to low light, is the standard flourescent laboratory assess the efficacy of changed housing conditions lighting too strong for them? A simple shield (Malham et al., 2002; Larson and Anderson, around the housing area might be all that is 2010). Alternatively, group-tolerant cuttlefish needed to approximate more ecologically may be a better alternative. For crustaceans that relevant conditions. Aside from a correct form loose dominance hierarchies based on day:night cycle, which is usually in place to retain resource holding potential (Gherardi and Daniels, appropriate circadian rhythms, other aspects of 2003; Fero et al., 2007; Irvin and Williams, 2009), lighting could be considered. If animals are providing more shelters than individuals in group startled by sudden bright light, the sudden lights- housing tanks can reduce stress and injury on in the morning might be improved by associated with territorial conflicts. installing a dimmer switch or timer that increases light levels slowly. Aquatic species, such as Promoting natural feeding behaviours freshwater crayfish and molluscs, may find bright Feeding methods also provide opportunities for lighting stressful or aversive (Cook and Carew, enrichment and otherwise improved welfare 1986; Barr and Elwood, 2011), a concern when outcomes. For animals that are normally we consider that chronically stressed animals predatory, is offering live an option? If live may yield quite different experimental results to food cannot be offered such that welfare of both those without such stressors. Other lighting predator and prey species is not compromised aspects such as spectrum, intensity and providing unreasonably (non-lethal injury of either, for light-sheltered areas may be worthwhile to example, is rare) or regular consumption rates consider. cannot be assured, is there another way of proving naturalistic foraging opportunity to the Stocking density of holding spaces should be animals? For cephalopods trained to accept dead based upon reasonable encounter rates in prey, frozen food can be suspended in water natural environments. Animals that school, currents to provide additional visual and olfactory swarm or form social groups should be permitted stimulation, which may enhance consumption to maintain similar structure if at all possible, rates (Hanley et al., 1998; Domingues et al., with this need balanced against the need to 2003). For terrestrial species, food that is buried individually identify members of a group; if or otherwise concealed behind barriers and must marking or labeling involves a stressful or be sought actively can simulate naturalistic potentially painful procedure, on balance foraging behaviours. For herbivores (e.g., individual housing may be more appropriate. locusts, marine and terrestrial snails), consider replacing or supplementing dried or artificial Are your animals normally solitary or territorial? diets with live plant material (Hinks and If so, group housing may be inappropriate even Erlandson, 1994; Capo et al., 2009), which most

14 Journal of Postdoctoral Research 2013: 9-20 likely better represents the nutritional profile the unfeasible or presents a considerable risk to animal requires (although extreme care should be experimenters (Dombrowski and De Voe, 2007; taken to avoid contaminated with Bennie et al., 2011; Harvey-Clark, 2011). pesticides or other chemicals). For all species, a varied diet is almost certainly better than a During transfer from one environment to homogenous one, and any opportunity to another, shielding the animal from unnecessary stimulate naturalistic foraging behaviours should visual and tactile disturbance should be a priority, be considered as worthwhile. even if the animal has limited sensory capacities in some modalities. Cover transfer containers if Experimental procedures possible, use careful handling to avoid jostling your animals about, and if animals are prone to Even more so than changing husbandry aggressive behaviours when stressed (crayfish procedures, making changes to experimental and squid, for example, will attack familiar procedures that have in the past yielded good conspecifics in transfer tanks when they had results can be challenging, even though previously cohabited in housing without improving animal welfare should not adversely concern), always transfer animals individually affect experimental outcomes. Experimental rather than in groups. procedures here are divided into two phases; restraint (involving handling, noninvasive Intervention examinations and noninvasive marking) and Intervention is considered any procedure that is intervention (covering invasive procedures, invasive (from injections to major surgery), and surgery, anesthesia, analgesia, and euthanasia). experimental procedures where the animal performs a task that is not part of its normal Restraint activity. Probably the most pressing and controversial issue related to intervention is pain: Restraint techniques should always aim to How can we tell if the animal experiences pain, protect both the experimenter and subject. Many and how can it be prevented? Because invertebrates have defensive weapons, some of invertebrate animals are phylogenetically, which present genuine dangers to experimenters. morphologically and behaviourally far removed Toxic and venomous animals should be handled from ourselves, these are difficult questions to with extreme care, but appropriate restraint that answer. reduce stress on the animal can also prevent deployment of defenses, improving outcomes for Anesthesia, analgesia and euthanasia everybody involved. Many of the same anesthetic agents appropriate for invertebrates are also effective for Procedures involving handling and immobility of euthanasia, although this is not universally true. invertebrates should be optimised to be as brief Analgesia is a more difficult topic, both because it as possible, but balanced against causing minimal is difficult to assess whether it might be disturbance and distress to the animal. Direct necessary (does the animal feel pain or is capable handling, although often faster, may be more only of nociceptive ?), and efficacy in stressful to the animal than secondary animals where pain is a valid concern is equally containment (allowing the animal to enter a difficult to discern. Maintenance analgesia is small, sealable transfer container, then moving rarely employed for invertebrates, and analgesia animal and container together). This technique is during procedures is typically accomplished by useful for arthropods such as stinging insects, either local or general anesthetics (Cooper, 2001, spiders and scorpions, venomous molluscs such 2011). as blue-ringed octopuses, stinging cnidarians, or any animal where acclimation to handling is Methods for euthanasia should be appropriate

Robyn J. Crook 15 for each animal’s physiology. The primary aim of immediate contraction at the decapitation site any euthanasia method is to kill the animal closes the wound: these factors suggest that the rapidly and without undue distress, while keeping squid’s may be functional for minutes after tissue in a state that is suitable for use in the decapitation. given experiment (AVMA, 2007). In some cases where unaltered physiology is particularly Of course, we do not know whether the animal important, anesthetic agents are not desirable suffers during this death; there is no evidence and a physical method of euthanasia is preferred. that cephalopods feel pain, despite their brains In all cases ensuring death has occurred prior to being highly complex (Crook and Walters, 2011; sample collection should be paramount. Many Crook et al., 2011). Nonetheless, this method of techniques that are reliable in vertebrate species killing can be greatly improved with very little are ineffective in invertebrates, so a means of additional time and effort. Simply decerebrating confirming death should be established. (using a scalpel blade to make 4-6 slices through the cranium, slicing the brain into multiple small A good example for improved euthanasia pieces) immediately after decapitating ensures methods can be found in the use of the squid that the brain is immediately dead, rather than Loligo pealei for studies on the giant axon. The allowing slow hypoxia to achieve brain death. giant axon and synapse have yielded many Death is clear from the immediately relaxation of fundamental insights into the functioning of the the chromatophores, giving a uniform pale nervous system, and are a common model for appearance, and the cessation of arm movement. studying action potential dynamics (Cole and This simple procedural change, which takes no Curtis, 1939; Hodgkin and Huxley, 1939, 1952; more than an extra 10 seconds, dramatically Hodgkin and Katz, 1949a, 1949b; Takeuchi and alters the death experience for squid used in such Takeuchi, 1962). The axon and its synapse are experiments. located in the mantle of the animal, so the chosen method of killing is decapitation, Because physical euthanasia typically requires removing the head at the point where it joins the close handling and experimenters must be mantle. This method is fast, requires minimal trained to ensure good technique, chemical training and does not require an anaesthetic euthanasia methods are preferable if they do not agent that might disrupt the physiology of the interfere with the intended use of the animal nervous system. However, anyone who has tissue. For aquatic species, bath immersion is a witnessed this preparation will be struck by the noninvasive method for delivering drugs, while continued, co-ordinated movements of the head- inhalants or injectables are typical for terrestrial end of the squid. The arms continue to move in species (Cooper, 1980, 2011). Whatever the sequence, are responsive to touch and the method used, animals should be monitored mantle-less animal will crawl about. The animal throughout the procedure and signs of distress appears to still be conscious; while animal bodies should indicate changes to the technique may be will make reflexive movements even after brain necessary. death, the coordinated nature of the squid movements and the attempts to escape suggest Isoflurane is a highly effective volatile anesthetic perhaps something more. The brain of the animal that is used on vertebrates and terrestrial is in the head, of course, and is not damaged by invertebrates alike. It is typically fast acting and decapitation. In contrast to vertebrates, there is readily reversible, and is used commonly for also a large tissue volume above the neck, anesthesia of terrestrial arthropods such as containing approximately 25% of the animals’ spiders (Cooper, 2011). However, its appropriate total blood volume. The hemostatic mechanism use requires specialized equipment and training, in molluscs is muscle contraction (Spurling, 1981; and its efficacy in humans after accidental Krontiris-Litowitz et al., 1989), meaning that exposure is well documented (Langley et al.,

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1995; Wenker, 1998; Epp and Waldner, 2012). Studies on invertebrate animal welfare; Carbon dioxide gas is effective for euthanasia informing regulation, disseminating the (but not analgesia) in most terrestrial message. invertebrates (Cooper, 2011), and is a cheap and effective means of euthanizing multiple animals In addition to efforts made directly at the level of simultaneously. individual laboratories, postdoctoral researchers and other young scientists can aim to improve Cold euthanasia (freezing) is a decreasingly animal welfare by making it a focus of their own common method of anesthetizing or killing studies. There is relatively little literature on invertebrates, as it has several notable welfare of invertebrate animal in research, and drawbacks. Cold may not produce plenty of scope for additions to it with careful antinociception in invertebrates, and is thus study. However, in a funding-poor environment inappropriate for surgical procedures (Zachariah, (where many of us find ourselves), extending or 2011). Primary euthanasia by freezing can make changing our research focus away from funded ascertaining death extremely difficult, as areas is risky, and the funding available for poikilothermic animals can withstand low welfare-based research on invertebrate animals temperatures by entering torpid or static states is minimal. Despite these limitations, if you have that are difficult to distinguish from death. made improvements in your lab practices or if Additionally, many vertebrate animals show improved welfare has improved your nociceptive responses to extreme cold, and experimental outcomes, publishing your homologous ion channels that transduce noxious observations could be of considerable personal cold are also found in some invertebrates, and scientific benefit – a brief communication to suggesting that cold may also be noxious for a welfare-focused journal, a poster at a invertebrates (Vriens et al., 2004). A primary goal conference, or simply mentioning your findings in of euthanasia should be to avoid sustained your talks are all relatively low-investment ways activation of nociceptive circuitry, thus cold of disseminating your own findings to other anesthesia/euthanasia may not meet this researchers. criterion. If you have invested time and effort into Irrespective of the technique used, ensuring about the naturalistic behaviours of your lab death has occurred within a reasonable animals, publishing an of normal and timeframe can be difficult. The use of typical aberrant behaviours observed under different endpoint indicators such as cessation of housing or experimental conditions would be a respiration, absence of reflexive responses to useful contribution, as such information can be strong stimulation, or cessation of heartbeat may difficult to find. If during characterization of not be accurate indicators of death. A secondary behaviour or physiology you find evidence for method of euthanasia should be used in nociception, affective states or something ambiguous cases. Once animals are insensible perhaps indicative of pain, a controlled (not just immobile; selecting the correct anesthesia or analgesia test might yield anesthetic agent is vital to ensure anesthesia and information valuable both for your own research not only paralysis), extensive tissue harvest, but also for the broader field of animal welfare. decerebration by pithing, removal of the brain or injection of chemicals directly to ganglia (Battison In talking with well-established investigators et al., 2000; Bennie et al., 2012), rapid freezing or using invertebrate models, I find that there is immersion into fixative or ethanol are all often an extensive base of unpublished acceptable methods for ensuring animals will not observations on behavior and physiology, which revive spontaneously (Cooper, 2011). may not be of particular relevance to their major research question but is nonetheless informative

Robyn J. Crook 17 for others focused specifically on the wellbeing of exclusion, too. research animals. While there is great value in improving the lives and deaths of the animals in A word about activist organisations our direct care, a far greater effect could be Scientific knowledge on invertebrate animal achieved by sharing our scattered observations welfare should not be shared only with other through publication in peer-reviewed journals. I scientists. and activist groups find most post-doctoral researchers are quite seeking a scientific view on issues of concern to strongly motivated by opportunities for them occasionally contact researchers working additional publications, so this is a mutualistic on aspects of welfare (although these are most goal. commonly about commercial harvest and food preparation rather than animal research). The For researchers whose experimental species is relationship between activists and scientists is either already, or likely to be, regulated, typically, and often justifiably, characterized as establishing a productive, open relationship with adversarial, but I nonetheless encourage your institutional vets and animal welfare colleagues to engage these queries if they can committee members should be a priority. If there provide an empirically supported view on issues are not established guidelines in place already for of welfare. While such interactions can be husbandry or procedures, find out if they have frustrating, they should be considered as voluntary standards or are willing to work with important as interactions with other scientists. you on establishing them. If regulations are likely Activists have a powerful and well-defined public to be imposed upon you in future, having a good presence, and their role in shaping past policies working relationship with your veterinary staff on animal welfare is unquestioned. In any effort and animal welfare committees will be to develop regulations of invertebrates in advantageous; in these cases it is the researchers research, activist organisations will almost who are typically far more familiar with the certainly have a role. However, certain biology of their particular species than committee organisations do target scientists using animals members and veterinary staff who may have very (and sometimes aim deliberately at graduate minimal previous exposure to invertebrate students and post-doctoral researchers), thus it is animals. Being included in discussions during essential to confer with your direct supervisors initial efforts to develop guidelines will almost and the public information office at your certainly mean a better outcome for researchers institution before responding to any such and animals alike. requests.

Finally, for those of us working with invertebrates Conclusions in currently unregulated research environments (in the US, for example), taking an active role in In the coming years it is likely that at least some future discussion of national regulation or invertebrate species in all major research legislation will be vital. Be proactive about countries will be included in legislation protecting establishing your message and position – which animals used in scientific research. In Europe, animals should be protected and on what Britain and Canada these changes are either grounds your opinion is based, what form already established or will take effect in the protections should take, and if there are animals coming several years. While the appetite for or procedures you feel confident should not be legislation in the US is probably likely to remain restricted (non-invasive or transiently invasive low, US researchers seeking to publish in journals procedures such as injections, or all procedures based in these regulated nations may find for animals whose nervous systems are clearly themselves asked to comply with rules from too simple to experience suffering or distress, for these nations for their work to appear in these example), be willing to advocate for their journals. Thus restrictions on invertebrate animal

18 Journal of Postdoctoral Research 2013: 9-20 research are likely to be felt broadly, even in shore , . Behavioural countries without their own, domestic legislation. Processes 86:340–344. Battison A, MacMillan R, MacKenzie A, Rose P, If we permit that some regulation is largely Cawthorn R, Horney B (2000) Use of injectable inevitable, it is young investigators at the start of potassium chloride for euthanasia of American their careers who therefore have the most lobsters (Homarus americanus). Comparative incentive to ensure that such regulation is Medicine 50:545–550. reasonable; well grounded scientifically and Baumans V (2005) Science-based assessment of appropriate to the biology of each species. Young animal welfare: laboratory animals. Revue scientists should thus make a concerted, Scientifique et Technique-Office International organized effort to establish their positions on des Epizooties 24:503. what constitutes acceptable practice. If we ignore Bennie M, Loaring C, Trim S (2011) Laboratory the opportunities we have now, to make changes husbandry of arboreal tarantulas to our own lab practices and to conduct research (Theraphosidae) and evaluation of to support our positions prior to further environmental enrichment. Animal restrictions being enacted, we are at risk of being Technology and Welfare 10:163. governed by restrictions implemented without Bennie NAC, Loaring CD, Bennie… MMG (2012) scientific grounding. An effective method for terrestrial arthropod euthanasia. The Journal of Experimental This is the real ‘danger’ of regulation of Biology. invertebrate use – that without a body of Bradley EA (1974) Some observations of Octopus evidence that we ourselves have gathered, any joubini reared in an inland aquarium. Journal new legislation will be poorly informed, with little of Zoology 173:355–368. relation to our animals’ biology and physiology. Capo TR, Bardales AT, Gillette PR, Lara MR, The science of invertebrate animal welfare is one Schmale MC, Serafy JE (2009) Larval growth, that that all young investigators should consider development, and survival of laboratory- as worthy and important, and only our continued reared californica: Effects of diet and non-engagement with these issues will ensure veliger density. Comparative Biochemistry and that we find ourselves bound by regulations Physiology Part C: Toxicology & Pharmacology within which we cannot work effectively. 149:215–223. Therefore I hope to see many more publications Carbone L (2004) What animals want: Expertise from post-docs, graduate students and young and advocacy in laboratory animal welfare faculty in the coming years on these important policy. Oxford University Press, USA. topics. Chase R (2002) Behavior and its neural control in gastropod molluscs. Oxford University Press, References USA. Cole KS, Curtis HJ (1939) Electric impedance of Adriaensen H, Gybels J, Handwerker HO, Van the squid giant axon during activity. The Hees J (1980) Latencies of chemically evoked Journal of General Physiology 22:649–670. discharges in human cutaneous nociceptors Cook DG, Carew TJ (1986) and of the concurrent subjective sensations. of head waving in Aplysia. Proceedings of the Letters 20:55–59. National Academy of Sciences 83:1120–1124. Allen C (2004) Animal pain. Noûs 38:617–643. Cooper JE (1980) Invertebrates and invertebrate AVMA AVMA (2007) Guidelines on euthanasia, disease; an introduction for the veterinary formerly report of the AVMA panel on surgeon. Journal of Small Animal Practice euthanasia. 20. 21:495–508. Barr S, Elwood RW (2011) No evidence of Cooper JE (2001) Invertebrate anesthesia. The morphine analgesia to noxious shock in the veterinary clinics of North America Exotic

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