Insects enter chill coma, a reversible state of paralysis, at temperatures below their critical thermal minimum (CT min ), and the time required for an insect to recover after a cold exposure is termed chill coma recovery time (CCRT). The CT min and CCRT are both important metrics of insect cold tolerance that are used interchangeably, although chill coma recovery is not necessarily permitted by a direct reversal of the mechanism causing chill coma onset. Nevertheless, onset and recovery of coma have been attributed to loss of neuromuscular function due to depolarization of muscle fibre membrane potential (V m does depolarize muscle resting potential and slow CCRT following prolonged cold exposure. However, onset of chill coma at the CT min relates to an as-yet-unknown effect of temperature on neuromuscular function.

Summary Chill tolerance of insects is defined as the ability of insects to tolerate low temperature under circumstances not involving freezing of intra- or extracellular fluids. For many insects chill tolerance is crucial for their ability to persist in cold environments and mounting evidence indicate that chill tolerance is associated with the ability to maintain ion- and water-homeostasis, thereby ensuring muscular function and preventing chill injury at low temperature. The present study describes the relationship between muscle and hemolymph ion-homeostasis and time to regain posture following cold shock (CS, 2h at -4°C) in the chill susceptible locust, Locusta migratoria. This relationship is examined in animals with and without a prior rapid cold hardening treatment (RCH, 2h at 0°C) to investigate the physiological underpinnings of RCH. Cold shock elicited a doubling of hemolymph [K+] and this disturbance was greater in locusts pre-exposed to RCH. Recovery of ion homeostasis was, however, markedly faster in RCH treated animals which correlated well with whole organism performance as hardened individuals regained posture more than 2 minutes faster than non-hardened individuals following CS. The present study indicates that loss and recovery of muscular function is associated with resting membrane potential of excitable membranes as attested from the changes in the equilibrium potential for K+ (EK) following CS. Both hardened and non-hardened animals recovered movement once K+ homeostasis was recovered to a fixed level (EK≈ -41 mV). RCH is therefore not associated with altered sensitivity to ion disturbance but instead a faster recovery of hemolymph [K+].

When exposed to low temperatures, many insect species enter a reversible comatose state (chill coma), which is driven by a failure of neuromuscular function. Chill coma and chill coma recovery have been associated with a loss and recovery of ion homeostasisand accordingly onset of chill coma has been hypothesized to result from depolarization of membrane potential caused by loss of ion homeostasis. Here, we examined whether onset of chill coma is associated with a disturbance in ion balance by examining the correlation between disruption of ion homeostasis and onset of chill coma in locusts exposed to cold at varying rates of cooling. Chill coma onset temperature changed maximally 1°C under different cooling rates and marked disturbances of ion homeostasis were not observed at any of the cooling rates. In a second set of experiments, we used isolated tibial muscle to determine how temperature and [K + ] o , independently and together, affect tetanic force production. Tetanic force decreased by 80% when temperature was reduced from 23°C to 0.5°C, while an increase in [K + ] o from 10 mmol l −1 to 30 mmol l −1 at 23°C caused a 40% reduction in force. Combining these two stressors almost abolished force production. Thus, low temperature alone may be responsible for chill coma entry, rather than a disruption of extracellular K + homeostasis. As [K + ] also has a large effect on tetanic force production, it is hypothesized that recovery of [K + ] o following chill coma could be important for the time to recovery of normal neuromuscular function.

SUMMARYNumerous recent studies convincingly correlate the upper thermal tolerance limit of aquatic ectothermic animals to reduced aerobic scope, and ascribe the decline in aerobic scope to failure of the cardiovascular system at high temperatures. In the present study we investigate whether this ʻaerobic scope modelʼ applies to an air-breathing and semi-terrestrial vertebrate Rhinella marina (formerly Bufo marinus). To quantify aerobic scope, we measured resting and maximal rate of oxygen consumption at temperatures ranging from 10 to 40°C. To include potential effects of acclimation, three groups of toads were acclimated chronically at 20, 25 and 30°C, respectively. The absolute difference between resting and maximal rate of oxygen consumption increased progressively with temperature and there was no significant decrease in aerobic scope, even at temperature immediately below the lethal limit (41-42 o C). Haematological and cardiorespiratory variables were measured at rest and immediately after maximal activity at benign (30°C) and critically high (40°C) temperatures. Within this temperature interval, both resting and active heart rate increased, and there was no indication of respiratory failure, judged from high arterial oxygen saturation, P O2 and [Hb O2 ]. With the exception of elevated resting metabolic rate for cold-acclimated toads, we found few differences in the thermal responses between acclimation groups with regard to the cardiometabolic parameters. In conclusion, we found no evidence for temperature-induced cardiorespiratory failure in R. marina, indicating that maintenance of aerobic scope and oxygen transport is unrelated to the upper thermal limit of this air-breathing semi-terrestrial vertebrate.

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