R. Meldrum Robertson

Activity of descending contralateral movement detector neurons and collision avoidance behaviour in response to head-on visual stimuli in locusts.

Abstract. We recorded the activity of the right and left descending contralateral movement detectors responding to 10-cm (small) or 20-cm (large) computer-generated spheres approaching along different trajectories in the locusts frontal field of view. In separate experiments we examined the steering responses of tethered flying locusts to identical stimuli. The descending contralateral movement detectors were more sensitive to variations in target trajectory in the horizontal plane than in the vertical plane. Descending contralateral movement detector activity was related to target trajectory and to target size and was most sensitive to small objects converging on a direct collision course from above and to one side. Small objects failed to induce collision avoidance manoeuvres whereas large objects produced reliable collision avoidance responses. Large targets approaching along a converging trajectory produced steering responses that were either away from or toward the side of approach of the object, whereas targets approaching along trajectories that were offset from the locusts mid-longitudinal body axis primarily evoked responses away from the target. We detected no differences in the discharge properties of the descending contralateral movement detector pair that could account for the different collision avoidance behaviours evoked by varying the target size and trajectories. We suggest that descending contralateral movement detector properties are better suited to predator evasion than collision avoidance.

Stress preconditioning of spreading depression in the locust CNS.

Cortical spreading depression (CSD) is closely associated with important pathologies including stroke, seizures and migraine. The mechanisms underlying SD in its various forms are still incompletely understood. Here we describe SD-like events in an invertebrate model, the ventilatory central pattern generator (CPG) of locusts. Using K+ -sensitive microelectrodes, we measured extracellular K+ concentration ([K+]o) in the metathoracic neuropile of the CPG while monitoring CPG output electromyographically from muscle 161 in the second abdominal segment to investigate the role K+ in failure of neural circuit operation induced by various stressors. Failure of ventilation in response to different stressors (hyperthermia, anoxia, ATP depletion, Na+/K+ ATPase impairment, K+ injection) was associated with a disturbance of CNS ion homeostasis that shares the characteristics of CSD and SD-like events in vertebrates. Hyperthermic failure was preconditioned by prior heat shock (3 h, 45°C) and induced-thermotolerance was associated with an increase in the rate of clearance of extracellular K+ that was not linked to changes in ATP levels or total Na+/K+ ATPase activity. Our findings suggest that SD-like events in locusts are adaptive to terminate neural network operation and conserve energy during stress and that they can be preconditioned by experience. We propose that they share mechanisms with CSD in mammals suggesting a common evolutionary origin.

Cloning and characterization of a member of the hsp70 gene family from Locusta migratoria, a highly thermotolerant insect

Abstract A complementary deoxyribonucleic acid (cDNA) and the corresponding gene segment encoding a member of the 70-kDa heat shock protein (Hsp70) family have been cloned and sequenced from Locusta migratoria, the African migratory locust. These animals are noted for their thermotolerance, which can exceed temperatures of 50°C. Conceptually translated, the sequence shows a 654-residue protein with theoretical molecular weight of 71.4 kDa, which more closely resembles the mammalian Hsp70 (84–85% similarity) than Hsp70 from other insects, with ∼75% similarity to the sequence from the fruit fly. Comparisons of cDNA and genomic sequences show that the gene contains 2 introns, a 245-bp intron located in the 5′ untranslated region and a 91-bp intron in the coding region. Transcript abundance, as estimated by Northern blot analysis and reverse transcription–polymerase chain reaction, shows that heat shock treatment (45°C for 3 hours) does not elevate hsp70 messenger ribonucleic acid levels in fat bodies or in neural tissues. Immunological assays of Hsp70 show that the protein is constitutively expressed, with a modest, ∼2-fold induction after a 3-hour heat shock in fat body preparations. Although this sequence could be an hsc70 rather than an hsp70, it was the only cDNA isolated from heat-shocked tissue. Whatever the formal designation, such modest induction and constitutive expression may be ideally suited as an adaptation to the locusts chronic exposure to heat shock temperatures and the consequent demand for chaperone proteins.

The Multiple Functions of Cysteine-String Protein Analyzed at Drosophila Nerve Terminals

The synaptic vesicle-associated cysteine-string protein (CSP) is important for synaptic transmission. Previous studies revealed multiple defects at neuromuscular junctions (NMJs) of csp null-mutant Drosophila, but whether these defects are independent of each other or mechanistically linked through J domain mediated-interactions with heat-shock cognate protein 70 (Hsc70) has not been established. To resolve this issue, we genetically dissected the individual functions of CSP by an in vivo structure/function analysis. Expression of mutant CSP lacking the J domain at csp null-mutant NMJs fully restored normal thermo-tolerance of evoked transmitter release but did not completely restore evoked release at room temperature and failed to reverse the abnormal intraterminal Ca2+ levels. This suggests that J domain-mediated functions are essential for the regulation of intraterminal Ca2+ levels but only partially required for regulating evoked release and not required for protecting evoked release against thermal stress. Hence, CSP can also act as an Hsc70-independent chaperone protecting evoked release from thermal stress. Expression of mutant CSP lacking the L domain restored neurotransmission and partially reversed the abnormal intraterminal Ca2+ levels, suggesting that the L domain is important, although not essential, for the role of CSP in regulating intraterminal Ca2+ levels. We detected no effects of csp mutations on individual presynaptic Ca2+ signals triggered by action potentials, suggesting that presynaptic Ca2+ entry is not primarily impaired. Both the J and L domains were also required for the role of CSP in synaptic growth. Together, these results suggest that CSP has several independent synaptic functions, affecting synaptic growth, evoked release, thermal protection of evoked release, and intraterminal Ca2+ levels at rest and during stimulation.

Coma in response to environmental stress in the locust: A model for cortical spreading depression

Spreading depression (SD) is an interesting and important phenomenon due to its role in mammalian pathologies such as migraine, seizures, and stroke. Until recently investigations of the mechanisms involved in SD have mostly utilized mammalian cortical tissue, however we have discovered that SD-like events occur in the CNS of an invertebrate model, Locusta migratoria. Locusts enter comas in response to stress during which neural and muscular systems shut down until the stress is removed, and this is believed to be an adaptive strategy to survive extreme environmental conditions. During stress-induced comas SD-like events occur in the locust metathoracic ganglion (MTG) that closely resemble cortical SD (CSD) in many respects, including mechanism of induction, extracellular potassium ion changes, and propagation in areas equivalent to mammalian grey matter. In this review we describe the generation of comas and the associated SD-like events in the locust, provide a description of the similarities to CSD, and show how they can be manipulated both by stress preconditioning and pharmacologically. We also suggest that locust SD-like events are adaptive by conserving energy and preventing cellular damage, and we provide a model for the mechanism of SD onset and recovery in the locust nervous system.

Suppression of Spreading Depression-Like Events in Locusts by Inhibition of the NO/cGMP/PKG Pathway

Despite considerable research attention focused on mechanisms underlying neural spreading depression (SD), because of its association with important human CNS pathologies, such as stroke and migraine, little attention has been given to explaining its occurrence and regulation in invertebrates. In the locust metathoracic ganglion (MTG), an SD-like event occurs during heat and anoxia stress, which results in cessation of neuronal output for the duration of the applied stress. SD-like events were characterized by an abrupt rise in extracellular potassium ion concentration ([K+]o) from a baseline concentration of ∼8 to >30 mm, which returned to near baseline concentrations after removal of the applied stress. After return to baseline [K+]o, neuronal output (ventilatory motor pattern activity) from the MTG recovered. Unlike mammalian neurons, which depolarize almost completely during SD, locust neurons only partially depolarized. SD-like events in the locust CNS were suppressed by pharmacological inhibition of the nitric oxide/cyclic guanosine monophosphate/protein kinase G (NO/cGMP/PKG) pathway and were exacerbated by its activation. Also, environmental stressors such as heat and anoxia increased production of nitric oxide in the locust CNS. Finally, for the intact animal, manipulation of the pathway affected the speed of recovery from suffocation by immersion under water. We propose that SD-like events in locusts provide an adaptive mechanism for surviving extreme environmental conditions. The highly conserved nature of the NO/cGMP/PKG signaling pathway suggests that it may be involved in modulating SD in other organisms, including mammals.

Natural Variation in the Thermotolerance of Neural Function and Behavior due to a cGMP-Dependent Protein Kinase

Although it is acknowledged that genetic variation contributes to individual differences in thermotolerance, the specific genes and pathways involved and how they are modulated by the environment remain poorly understood. We link natural variation in the thermotolerance of neural function and behavior in Drosophila melanogaster to the foraging gene (for, which encodes a cGMP-dependent protein kinase (PKG)) as well as to its downstream target, protein phosphatase 2A (PP2A). Genetic and pharmacological manipulations revealed that reduced PKG (or PP2A) activity caused increased thermotolerance of synaptic transmission at the larval neuromuscular junction. Like synaptic transmission, feeding movements were preserved at higher temperatures in larvae with lower PKG levels. In a comparative assay, pharmacological manipulations altering thermotolerance in a central circuit of Locusta migratoria demonstrated conservation of this neuroprotective pathway. In this circuit, either the inhibition of PKG or PP2A induced robust thermotolerance of neural function. We suggest that PKG and therefore the polymorphism associated with the allelic variation in for may provide populations with natural variation in heat stress tolerance. fors function in behavior is conserved across most organisms, including ants, bees, nematodes, and mammals. PKGs role in thermotolerance may also apply to these and other species. Natural variation in thermotolerance arising from genes involved in the PKG pathway could impact the evolution of thermotolerance in natural populations.

Neuronal circuits controlling flight in the locust: central generation of the rhythm

Abstract How is the nervous system organized to produce the complex motor patterns which underlie behaviour? Can behaviour be adequately described in terms of the properties and interactions of single neurons? These questions have been addressed in a number of motor systems and some progress has been made in describing central neuronal mechanisms generating motor patterns (e.g. Ref. 1). However, behaviour involves an interaction of an animal with a changing environment and thus sensory as well as central processes must be investigated before complete success in answering such questions can be claimed. Locust flight lends itself particulary well to both such investigations (e.g. Ref. 2) and this system may prove to be the most tractable for a cellular analysis of behaviour. In this article I review recent results concerning the central generation of the flight motor pattern of the locust 3–5 and in the following article Reichert and Rowell review their advances in understanding of how information from the periphery modifies this centrally generated motor pattern 6,7 .

Temperature and neuronal circuit function: compensation, tuning and tolerance

Temperature has widespread and diverse effects on different subcellular components of neuronal circuits making it difficult to predict precisely the overall influence on output. Increases in temperature generally increase the output rate in either an exponential or a linear manner. Circuits with a slow output tend to respond exponentially with relatively high Q(10)s, whereas those with faster outputs tend to respond in a linear fashion with relatively low temperature coefficients. Different attributes of the circuit output can be compensated by virtue of opposing processes with similar temperature coefficients. At the extremes of the temperature range, differences in the temperature coefficients of circuit mechanisms cannot be compensated and the circuit fails, often with a reversible loss of ion homeostasis. Prior experience of temperature extremes activates conserved processes of phenotypic plasticity that tune neuronal circuits to be better able to withstand the effects of temperature and to recover more rapidly from failure.

Modulation of Neural Circuit Operation by Prior Environmental Stress

Abstract Many organisms are exposed to harsh environmental conditions that may impair the operation of vital neuronal circuits and imperil the animal before these conditions directly cause cell and tissue death. Prior exposure to extreme but sub-lethal stress has long-term effects on neural circuit function enabling motor pattern generators to operate under previously non-permissive conditions. Using several model systems we have been investigating the mechanisms underlying stress-mediated neuroprotection, particularly thermotolerance imparted by a prior heat shock. Prior anoxia and cold shock also impart thermotolerance of motor pattern generation suggesting that different stressors activate common protective pathways. Synaptic transmission, action potential generation and neuronal potassium conductance are modulated by prior heat shock. Pharmacological block of potassium channels, which increases the duration of action potentials and the amplitude of postsynaptic potentials, mimics the thermoprotective effect of a prior heat shock. A universal consequence of heat shock and other stresses is the increased expression of a suite of heat shock proteins of which HSP70 is most closely linked to organismal thermotolerance. Increased levels of HSP70 are sufficient, but not necessary for synaptic thermoprotection. Accumulating evidence suggests the existence of multiple, overlapping pathways for protection and that these mechanisms may be neuron specific depending on their functional roles.