Stress-Induced Antinociception in Fish Reversed by Naloxone

Abstract

Pain perception in non-mammalian vertebrates such as fish is a controversial issue. We demonstrate that, in the fish Leporinus macrocephalus, an imposed restraint can modulate the behavioral response to a noxious stimulus, specifically the subcutaneous injection of 3% formaldehyde. In the first experiment, formaldehyde was applied immediately after 3 or 5 min of the restraint. Inhibition of the increase in locomotor activity in response to formaldehyde was observed, which suggests a possible restraint-induced antinociception. In the second experiment, the noxious stimulus was applied 0, 5, 10 and 15 min after the restraint, and both 3 and 5 min of restraint promoted short-term antinociception of approximately 5 min. In experiments 3 and 4, an intraperitoneal injection of naloxone (30 mg.kg−1) was administered 30 min prior to the restraint. The 3- minute restraint-induced antinociception was blocked by pretreatment with naloxone, but the corresponding 5-minute response was not. One possible explanation for this result is that an opioid and a non-preferential μ–opioid and/or non-opioid mechanism participate in this response modulation. Furthermore, we observed that both the 3- and 5- minutes restraint were severely stressful events for the organism, promoting marked increases in serum cortisol levels. These data indicate that the response to a noxious stimulus can be modulated by an environmental stressor in fish, as is the case in mammals. To our knowledge, this study is the first evidence for the existence of an endogenous antinociceptive system that is activated by an acute standardized stress in fish. Additionally, it characterizes the antinociceptive response induced by stress in terms of its time course and the opioid mediation, providing information for understanding the evolution of nociception modulation.

Acute administration of a cannabinoid CB1 receptor antagonist impairs stress-induced antinociception in fish

“…Subcutaneous injections of 3% formaldehyde in the region of the adipose fin were used as the noxious stimulus, reproducing the nociceptive models previously used in rats [19] and adapted for fish in our laboratory [3,4]. The animals were removed from the water with nylon nets, wrapped in wet cloth for formaldehyde injection, and immediately returned to the water.…”

“…The procedure was performed without anaesthesia to allow evaluation of the initial nociceptive responses and was as brief as possible (approximately 5 s) to minimize suffocation. The subcutaneous injection of 3% formaldehyde, but not saline, promotes an immediate and intense increase in locomotor activity, at levels above the normal pattern for the species, associated with an erratic swim pattern [3,4], which was used as an indicator of nociception.…”

“…The restraint stress was used to induce antinociception in L. macrocephalus, as described by Wolkers et al [4]. In this species the application of restraint stress prior the 3% formaldehyde subcutaneous injection can inhibit the increase in locomotor activity promoted by this noxious stimulation.…”

“…In Leporinus macrocephalus, the systemic pre-treatment with naloxone, a non-selective opioid antagonist, blocked the antinociception induced by conspecific alarm substance [3] and 3 min of restraint stress [4]. However, the naloxone systemic pre-treatment did not have the same effect on the antinociception induced by 5 min of restraint stress, suggesting that the modulation of restraint-induced antinociception in fish can depend on temporal parameters of the restraint and can involve other neurochemical systems in addition to opioid mechanisms [4].…”

“…Although the stress-induced antinociceptive systems are well known in mammals, the knowledge in fish is still limited. Recent studies have demonstrated that different sorts of external stressors, such as social subordination [2], conspecific alarm substance [3] and restraint stress [4], can promote an inhibition of nociceptive responses in fish, suggesting the activation of an endogenous antinociceptive system.…”