The Ionic Requirements for the Initiation of Action Potentials in Insect Muscle Fibers

SUMMARY In the hive, a wide range of honeybees tasks such as cell cleaning,nursing, thermogenesis, flight, foraging and inter-individual communication(waggle dance, antennal contact and trophallaxy) depend on proper muscle activity. However, whereas extensive electrophysiological studies have been undertaken over the past ten years to characterize ionic currents underlying the physiological neuronal activity in honeybee, ionic currents underlying skeletal muscle fibre activity in this insect remain, so far, unexplored. Here, we show that, in contrast to many other insect species, action potentials in muscle fibres isolated from adult honeybee metathoracic tibia,are not graded but actual all-or-none responses. Action potentials are blocked by Cd2+ and La3+ but not by tetrodotoxin (TTX) in current-clamp mode of the patch-clamp technique, and as assessed under voltage-clamp, both Ca2+ and K+ currents are involved in shaping action potentials in single muscle fibres. The activation threshold potential for the voltage-dependent Ca2+ current is close to–40 mV, its mean maximal amplitude is –8.5±1.9 A/F and the mean apparent reversal potential is near +40 mV. In honeybees, GABA does not activate any ionic membrane currents in muscle fibres from the tibia, but L-glutamate, an excitatory neurotransmitter at the neuromuscular synapse induces fast activation of an inward current when the membrane potential is voltage clamped close to its resting value. Instead of undergoing desensitization as is the case in many other preparations, a component of this glutamate-activated current has a sustained component, the reversal potential of which is close to 0 mV, as demonstrated with voltage ramps. Future investigations will allow extensive pharmacological characterization of membrane ionic currents and excitation–contraction coupling in skeletal muscle from honeybee, a useful insect that became a model to study many physiological phenomena and which plays a major role in plant pollination and in stability of environmental vegetal biodiversity.

Section: Discussionsupporting

confidence: 91%

Section: Introductionmentioning

confidence: 94%

Excitable properties of adult skeletal muscle fibres from the honeybeeApis mellifera

Collet

Belzunces

2007

“…Because some of the cells are excited through the nerves, any change in force production could in principle relate to changes in excitation and propagation of neuronal action potentials. To test this possibility, we performed experiments in the presence and absence of TTX, which is a potent blocker of nervous function but has no effect on locust muscle (Washio, 1972;Orchard et al, 1981;Collet, 2009). Application of TTX reduced overall force production to about half that in untreated muscle, indicating that approximately half of the muscle cells were exclusively dependent on nervous signal transmission (Fig.…”

Section: Mechanisms Behind Recovery Of Muscle Function Following Chilmentioning

confidence: 99%

“…Firstly, a force-frequency relationship was performed at both 23°C and 0.5°C to ensure that maximum force was reached when stimulating at 60 Hz for 2 s. Secondly, the stability of the preparation was examined to investigate whether muscle force production changed over time. This was performed on preparations with and without TTX (which blocks nervous function but has no effect on locust muscle) (Washio, 1972;Orchard et al, 1981) to further test whether a reduction in force with time was associated with reduced muscle function or failure in the excitation of associated motor neurons.…”

Section: Control Series Of Force Measurementsmentioning

confidence: 99%

Why do insects enter and recover from chill coma? Low temperature and high extracellular potassium compromise muscle function inLocusta migratoria

Findsen

Pedersen

Petersen

et al. 2014

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.

“…In insect skeletal muscle, E-C coupling starts when an AP reaches the presynaptic terminal and triggers release of glutamate into the synaptic cleft (Kerkut et al, 1965;Usherwood and Machili, 1966). Glutamate binds to receptors in the post-synaptic muscle fiber membrane, resulting in a net inward movement of cations ( primarily Na + ), creating an endplate potential (EPP) in the muscle fiber (Washio, 1972;Salkoff and Wyman, 1983;Collet and Belzunces, 2007). If the EPP is sufficiently large, an AP is formed, driven by current through L-type voltage-gated Ca 2+ channels (Frolov and Singh, 2013).…”

Section: Introductionmentioning

confidence: 99%

Reduced L-type Ca2+ current and compromised excitability induce loss of skeletal muscle function during acute cooling in locust

Findsen

Overgaard

Pedersen

2016

Low temperature causes most insects to enter a state of neuromuscular paralysis, termed chill coma. The susceptibility of insect species to chill coma is tightly correlated to their distribution limits and for this reason it is important to understand the cellular processes that underlie chill coma. It is known that muscle function is markedly depressed at low temperature and this suggests that chill coma is partly caused by impairment in the muscle per se. To find the cellular mechanism(s) underlying muscle dysfunction at low temperature, we examined the effect of low temperature (5°C) on several events in excitation-contraction coupling in the migratory locust (Locusta migratoria). Intracellular membrane potential recordings during single nerve stimulations showed that 70% of fibers at 20°C produced an action potential (AP), while only 55% of fibers were able to fire an AP at 5°C. Reduced excitability at low temperature was caused by an ∼80% drop in L-type Ca 2+ current and a depolarizing shift in its activation of around 20 mV, which means that a larger endplate potential would be needed to activate the muscle AP at low temperature. In accordance, we showed that intracellular Ca 2+ transients were largely absent at low temperature following nerve stimulation. In contrast, maximum contractile force was unaffected by low temperature in chemically skinned muscle bundles, which demonstrates that the function of the contractile filaments is preserved at low temperature. These findings demonstrate that reduced L-type Ca 2+ current is likely to be the most important factor contributing to loss of muscle function at low temperature in locust.