Biomedical subjects

R Meldrum Robertson

Publications and source records attributed to R Meldrum Robertson.

14 recordsLinked to original sources

PubMed2006

Octopamine mediates thermal preconditioning of the locust ventilatory central pattern generator via a cAMP/protein kinase A signaling pathway.

We investigated the role of biogenic amines in generating thermoprotection of the ventilatory motor pattern circuitry in Locusta migratoria. Levels of octopamine (OA) and dopamine (DA) in the metathoracic ganglion decreased during heat stress. We measured the thermosensitivity of central pattern generation in response to a ramped increase of temperature in semi-intact preparations. OA, DA, and tyramine (TA) were either bath applied or injected into the locust hemocoel 4-8 h before testing. Neither TA nor DA modified the thermotolerance of ventilatory motor pattern generation. However, OA treatment by bath applications (10(-4) M OA) or by injections into the hemocoel (2 microg/10 microl OA) mimicked heat shock preconditioning and improved the thermotolerance of the motor pattern by increasing the failure temperature and by decreasing the time taken to recover operation after a return to room temperature. Heat shock-induced thermoprotection was eradicated in locusts preinjected with epinastine (Oct betaR antagonist). Neuropil injections of the cAMP agonist and protein kinase A (PKA) activator, Sp-cAMPs, both conferred thermoprotection in control locusts and rescued thermoprotection in epinastine-treated HS locusts. Similar injections of the PKA inhibitor Rp-cAMPs blocked the thermoprotective effect of bath-applied OA. Octopamine-mediated thermoprotection was also abolished with neuropil injections of cycloheximide or actinomycin D, indicating a requirement for transcription and translation. We conclude that OA has a crucial role in triggering protein synthesis-dependent physiological adaptations to protect CNS function during heat stress by activating a cAMP/PKA pathway.

Action Potentials↗

PubMed2006

Temperature-sensitive gating in a descending visual interneuron, DCMD.

Activity in neural circuits can be modified through experience-dependent mechanisms. The effects of high temperature on a locust visual interneuron (the descending contralateral movement detector, DCMD) have previously been shown to be mitigated by prior exposure to sub-lethal, elevated temperatures (heat shock, HS). Activity in the DCMD is reduced at high temperature in naïve animals (control), whereas HS animals show a maintained spike count at all temperatures. We examined whether this finding was due to direct effects of temperature on visual processing, or whether other indirect feedback mechanisms were responsible for the observed effect in the DCMD. Activity in the DCMD was elicited using a computer-generated looming image, and the response was recorded extracellularly. The temperature of visual processing circuits contributes directly to HS-induced plasticity in the DCMD, as maintaining the brain at 25 degrees C during a thoracic temperature ramp eliminated the high frequency activity associated with HS. Removing ascending input by severing the thoracic nerve cord reduced DCMD thermosensitivity, indicating that indirect feedback mechanisms are also involved in controlling the DCMD response to increased thoracic temperature. Understanding how thermosensitive feedback within the locust affects DCMD function provides insight into critical regulatory mechanisms underlying visually-guided behaviors.

Adaptation, Physiological↗

PubMed2006

Photoperiod-induced plasticity of thermosensitivity and acquired thermotolerance in Locusta migratoria.

The mechanisms by which different life histories affect neural circuits are largely unknown. We show that the thermosensitivity and thermotolerance of neural circuit operation are affected in a complex dynamic fashion by photoperiod, prior heat experience and the sex of the animal. We compared thermosensitivity and thermotolerance of ventilatory motor pattern generation in locusts reared under two photoperiods (12:12 and 16:8; i.e. 12 h:12 h and 16 h:8 h L:D, respectively) before and after heat shock pre-treatment (HS: 3 h, 45 degrees C) in order to determine the effect of daylength on properties of neural function. We monitored central pattern generator (CPG) output electromyographically from muscle 161 in the second abdominal segment during ramped increases in temperature and also measured the time taken for the circuit to fail at high temperatures and the time taken to recover on return to room temperature. There were effects of photoperiod, heat pre-treatment and the sex of the animal on ventilatory rate, time-to-failure and time-to-recovery. The ventilatory motor pattern of 16:8 and 12:12 locusts responded differently to increasing and maintained high temperature stress in both control and heat shocked locusts. We found that 12:12 locusts were generally more robust than 16:8 locusts: they lived longer, they showed greater tolerance to high temperatures, and they recovered more quickly from temperature-induced circuit failure. A faster ventilatory rate in 12:12 animals at high temperatures may have accelerated evaporative cooling to mediate improved temperature tolerance.

Animals↗

PubMed2005

Heat shock-mediated thermoprotection of larval locomotion compromised by ubiquitous overexpression of Hsp70 in Drosophila melanogaster.

Maintaining the competence of locomotor circuitry under stressful conditions can benefit organisms by enabling locomotion to more tolerable microhabitats. We show that prior heat shock protects locomotion and the locomotor central pattern generator of larval Drosophila against subsequent hyperthermic stress. We combined molecular genetic, electrophysiological, and behavioral techniques to investigate heat shock-mediated thermoprotection. Prior heat shock increased the distance traveled by larvae during hyperthermia before failure. The frequency of the rhythm of peristaltic locomotor contractions and the velocity of locomotion were both less thermosensitive after heat shock and were less susceptible to failure at high temperatures. Rhythmic coordinated motor patterns, recorded intracellularly as excitatory junction potentials in body wall muscles of dissected preparations, were centrally generated because patterns could still be generated in the absence of sensory feedback (sensory function disrupted with shibire). Prior heat shock protected central circuit operation during hyperthermic stress by increasing the temperature at which it failed. Overexpression of Hsp70 after a heat shock using transgenic flies (traII) did not enhance thermoprotection, as expected, but had deleterious effects on parameters of behavior.

Adaptation, Physiological↗

PubMed2005

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.

Analysis of Variance↗

PubMed2005

Synaptic thermoprotection in a desert-dwelling Drosophila species.

Synaptic transmission is a critical mechanism for transferring information from the nervous system to the body. Environmental stress, such as extreme temperature, can disrupt synaptic transmission and result in death. Previous work on larval Drosophila has shown that prior heat-shock exposure protects synaptic transmission against failure during subsequent thermal stress. This induced thermoprotection has been ascribed to an up-regulation of the inducible heat-shock protein, Hsp70. However, the mechanisms mediating natural thermoprotection in the wild are unknown. We compared synaptic thermosensitivity between D. melanogaster and a desert species, D. arizonae. Synaptic thermosensitivity and the functional limits of the related locomotor behavior differed significantly between closely related, albeit ecologically distinct species. Locomotory behavior of wandering third instar D. arizonae larvae was less thermosensitive and the upper temperature limit of locomotory function exceeded that of D. melanogaster by 6 degrees C. Behavioral results corresponded with significantly lower synaptic thermosensitivity at the neuromuscular junction in D. arizonae. Prior heat-shock protected only D. melanogaster by increasing relative excitatory junctional potential (EJP) duration, the time required for EJP failure at 40 degrees C, and the incidence of EJP recovery following heat-induced failure. Hsp70 induction profiles following heat-shock demonstrate up-regulation of inducible Hsp70 in D. melanogaster but not in D. arizonae. However, expression of Hsp70 under control conditions is greater in D. arizonae. These results suggest that the mechanisms of natural thermoprotection involve an increase in baseline Hsp70 expression.

Animals↗

PubMed2004

Heat stress-mediated plasticity in a locust looming-sensitive visual interneuron.

Neural circuits are strongly affected by temperature and failure ensues at extremes. However, detrimental effects of high temperature on neural pathways can be mitigated by prior exposure to high, but sublethal temperatures (heat shock). Using the migratory locust, Locusta migratoria, we investigated the effects of heat shock on the thermosensitivity of a visual interneuron [the descending contralateral movement detector (DCMD)]. Activity in the DCMD was elicited using a looming stimulus and the response was recorded from the axon using intracellular and extracellular methods. The thoracic region was perfused with temperature-controlled saline and measurements were taken at 5 degrees intervals starting at 25 degrees C. Activity in DCMD was decreased in control animals with increased temperature, whereas heat-shocked animals had a potentiated response such that the peak firing frequency was increased. Significant differences were also found in the thermosensitivity of the action potential properties between control and heat-shocked animals. Heat shock also had a potentiating effect on the amplitude of the afterdepolarization. The concurrent increase in peak firing frequency and maintenance of action potential properties after heat shock could enhance the reliability with which DCMD initiates visually guided behaviors at high temperature.

Action Potentials↗

PubMed2004

A role for the cytoskeleton in heat-shock-mediated thermoprotection of locust neuromuscular junctions.

A prior hyperthermic stress (heat shock) can induce thermoprotection of neuromuscular transmission in Locusta migratoria extensor tibiae muscle measured 4 h after the onset of the heat shock. It is not clear what effect an acute hyperthermic stress may have on the nervous system's ability to tolerate thermal stress, that is, before increased expression of heat-shock proteins. We found that over consecutive thermal stress tests, failure temperature was not altered in either heat-shock or control animals. This suggests that protective mechanisms are not established in the short term (within one hour). Various members of the heat-shock protein family interact with elements of the cytoskeleton. We found that preexposure of the preparation to cytoskeletal stabilizing drugs induced thermoprotection, while preexposure to cytoskeletal disrupting drugs disrupted the ability to confer and maintain thermoprotection. We conclude that thermoprotection relies on a stable cytoskeleton and suggest that members of the heat shock protein family are involved.

Animals↗

PubMed2003

Stress-induced thermotolerance of ventilatory motor pattern generation in the locust, Locusta migratoria.

Ventilation is a crucial motor activity that provides organisms with an adequate circulation of respiratory gases. For animals that exist in harsh environments, an important goal is to protect ventilation under extreme conditions. Heat shock, anoxia, and cold shock are environmental stresses that have previously been shown to trigger protective responses. We used the locust to examine stress-induced thermotolerance by monitoring the ability of the central nervous system to generate ventilatory motor patterns during a subsequent heat exposure. Preparations from pre-stressed animals had an increased incidence of motor pattern recovery following heat-induced failure, however, prior stress did not alter the characteristics of the ventilatory motor pattern. During constant heat exposure at sub-lethal temperatures, we observed a protective effect of heat shock pre-treatment. Serotonin application had similar effects on motor patterns when compared to prior heat shock. These studies are consistent with previous studies that indicate prior exposure to extreme temperatures and hypoxia can protect neural operation against high temperature stress. They further suggest that the protective mechanism is a time-dependent process best revealed during prolonged exposure to extreme temperatures and is mediated by a neuromodulator such as serotonin.

Adaptation, Physiological↗

PubMed2003

Cloning and characterization of a member of the Hsp70 gene family from Locusta migratoria, a highly thermotolerant insect.

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 degrees 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 approximately 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 degrees 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, approximately 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 locust's chronic exposure to heat shock temperatures and the consequent demand for chaperone proteins.

Amino Acid Sequence↗

PubMed2002

Impairment of central pattern generation in Drosophila cysteine string protein mutants.

The cysteine string proteins are integral synaptic vesicle proteins, critical for fast, Ca2+ -regulated exocytosis. Drosophila larvae with null mutations for the cysteine string protein (csp) gene have temperature-sensitive impairments of neurotransmission and presynaptic calcium removal at the neuromuscular junction. Using the larval Drosophila preparation to examine central pattern generation, we characterized the temperature sensitivity of locomotor patterns in wildtype and csp mutant larvae. Intraburst frequency of motoneuronal activity reached 100 Hz and was sufficiently high to rescue the temperature-sensitive synaptic failure in the mutant. Nevertheless, we show that deletion of the csp gene resulted in a severe deficiency in the generation of coordinated larval motor rhythms. Csp mutants that could generate patterned motor activity had slower, poorly coordinated rhythms with altered temperature sensitivity. We conclude that the temperature-sensitive paralysis characteristic of csp mutants is not a direct result of synaptic failure at neuromuscular junctions, as might be expected, but is the result of a failure of locomotor circuit operation at a higher integrative level.

Action Potentials↗

PubMed2002

Thermal preconditioning and heat-shock protein 72 preserve synaptic transmission during thermal stress.

As with other tissues, exposing the mammalian CNS to nonlethal heat stress (i.e., thermal preconditioning) increases levels of heat-shock proteins (Hsps) such as Hsp70 and enhances the viability of neurons under subsequent stress. Using a medullary slice preparation from a neonatal mouse, including the site of the neural network that generates respiratory rhythm (the pre-Bötzinger complex), we show that thermal preconditioning has an additional fundamental effect, protection of synaptic function. Relative to 30 degrees C baseline, initial thermal stress (40 degrees C) greatly increased the frequency of synaptic currents recorded without pharmacological manipulation by approximately 17-fold (p < 0.01) and of miniature postsynaptic currents (mPSCs) elicited by GABA (20-fold) glutamate (10-fold), and glycine (36-fold). Thermal preconditioning (15 min at 40 degrees C) eliminated the increase in frequency of overall synaptic transmission during acute thermal stress and greatly attenuated the frequency increases of GABAergic, glutamatergic, and glycinergic mPSCs (for each, p < 0.05). Moreover, without thermal preconditioning, incubation of slices in solution containing inducible Hsp70 (Hsp72) mimicked the effect of thermal preconditioning on the stress-induced release of neurotransmitter. That preconditioning and exogenous Hsp72 can affect and preserve normal physiological function has important therapeutic implications.

Animals↗

PubMed2002

Enhancement of presynaptic performance in transgenic Drosophila overexpressing heat shock protein Hsp70.

Prior heat shock confers protection to Drosophila synapses during subsequent heat stress by stabilizing quantal size and reducing the decline of quantal emission at individual synaptic boutons. The major heat shock protein Hsp70, which is strongly induced by high temperatures in Drosophila, may be responsible for this synaptic protection. To test this hypothesis, we investigated synaptic protection and stabilization at larval neuromuscular junctions of transgenic Drosophila which produce more than the normal amount of Hsp70 in response to heat shock. Overexpression of Hsp70 coincides with enhanced protection of presynaptic performance, assayed by measuring mean quantal content and percentage success of transmission. Quantal size was not selectively altered, indicating no effects of overexpression on postsynaptic performance. Thus, presynaptic mechanisms can be protected by manipulating levels of Hsp70, which would provide stability to neural circuits otherwise susceptible to heat stress.

Animals↗

PubMed1998

Heat shock protects synaptic transmission in flight motor circuitry of locusts.

We investigated the effects of heat shock on the temperature sensitivity of synaptic transmission in the motor circuit for flight in Locusta migratoria. In heat shocked animals synaptic transmission failed at 5-6 degrees C higher than in control animals and recovery of transmission was more than three times faster upon return to room temperature. We also found that synaptic delay was rendered insensitive to increases in temperature by heat shock. Thus we have shown in the locust that heat shock has important protective effects on synaptic transmission, thereby extending the upper temperature limit for the motor patterns that generate flight. This is the first description of an effect of heat shock that preserves neuronal communication under subsequent stressful conditions.

Animals↗