Päivi H. Torkkeli

Frequency response functions and information capacities of paired spider mechanoreceptor neurons

Abstract. Pseudorandom white-noise stimulation followed by direct spectral estimation was used to obtain linear frequency response and coherence functions from paired, but dynamically different, spider mechanosensory neurons. The dynamic properties of the two neuron types were similar with either mechanical or electrical stimulation, showing that action potential encoding dominates the dynamics. Phase-lag data indicated that action potential initiation occurs more rapidly during mechanical stimulation, probably in the distal sensory dendrites. Total information capacity, calculated from coherence, as well as information per action potential, were both similar in the two types of neurons, and similar to the few available estimates from other spiking neurons. However, information capacity and information per action potential both depended strongly on neuronal firing rate, which has not been reported before.

From stress and strain to spikes: mechanotransduction in spider slit sensilla

Abstract. This review focuses on the structure and function of a single mechanoreceptor organ in the cuticle of spiders. Knowledge emerging from the study of this organ promises to yield general principles that can be applied to mechanosensation in a wide range of animal systems. The lyriform slit sense organ on the antero-lateral leg patella of the spider Cupiennius salei is unusual in possessing large sensory neurons, whose cell bodies are close to the sites of sensory transduction, and accessible to intracellular recording during mechanotransduction. This situation, combined with recent technical developments, has made it possible to observe and experiment with all the major stages of mechanosensation. Important findings include the approximate size, number and ionic selectivity of the ion channels responsible for mechanotransduction, the types of voltage-activated ion channels responsible for action potential encoding, and the mechanisms controlling the dynamic properties of transduction and encoding. Most recently, a complex efferent system for peripheral modulation of mechanosensation has been discovered and partially characterized. Much remains to be learned about mechanosensation, but the lyriform slit sense organ system continues to offer important opportunities to advance our understanding of this crucial sense.

Spider Peripheral Mechanosensory Neurons Are Directly Innervated and Modulated by Octopaminergic Efferents

Octopamine is a chemical relative of noradrenaline providing analogous neurohumoral control of diverse invertebrate physiological processes. There is also evidence for direct octopaminergic innervation of some insect peripheral tissues. Here, we show that spider peripheral mechanoreceptors are innervated by octopamine-containing efferents. The mechanosensory neurons have octopamine receptors colocalized with synapsin labeling in the efferent fibers. In addition, octopamine enhances the electrical response of the sensory neurons to mechanical stimulation. Spider peripheral mechanosensilla receive extensive efferent innervation. Many efferent fibers in the legs of Cupiennius salei are GABAergic, providing inhibitory control of sensory neurons, but there is also evidence for other neurotransmitters. We used antibody labeling to show that some efferents contain octopamine and that octopamine receptors are concentrated on the axon hillocks and proximal soma regions of all mechanosensory neurons in the spider leg. Synaptic vesicles in efferent neurons were concentrated in similar areas. Octopamine, or its precursor tyramine, increased responses of mechanically stimulated filiform (trichobothria) leg hairs. This effect was blocked by the octopamine antagonist phentolamine. The octopamine-induced modulation was mimicked by 8-Br-cAMP, a cAMP analog, and blocked by Rp-cAMPS, a protein kinase A inhibitor, indicating that spider octopamine receptors activate adenylate cyclase and increase cAMP concentration. Frequency response analysis showed that octopamine increased the sensitivity of the trichobothria neurons over a broad frequency range. Thus, the major effect of octopamine is to increase its overall sensitivity to wind-borne signals from sources such as flying insect prey or predators.

Peripheral GABAergic inhibition of spider mechanosensory afferents

Spider mechanosensory neurons receive an extensive network of efferent synapses onto their sensory dendrites, somata and distal axonal regions. The function of these synapses is unknown. Peripheral synapses are also found on crustacean stretch‐receptor neurons but not on mechanosensory afferents of other species, although inhibitory GABAergic synapses are a common feature of centrally located axon terminals. Here we investigated the effects of GABA receptor agonists and antagonists on one group of spider mechanosensory neurons, the slit sense organ VS‐3, which are accessible to current‐ and voltage‐clamp recordings. Bath application of GABA activated an inward current that depolarized the membrane and increased the membrane conductance leading to impulse inhibition. VS‐3 neuron GABA receptors were activated by muscimol and inhibited by picrotoxin but not bicuculline, and their dose–response relationship had an EC50 of 103.4 µm, features typical for insect ionotropic GABA receptors. Voltage‐ and current‐clamp analysis confirmed that, while the Na+ channel inhibition resulting from depolarization can lead to impulse inhibition, the increase in membrane conductance (i.e. ‘shunting’) completely inhibited impulse propagation. This result argues against previous findings from other preparations that GABA‐mediated inhibition is caused by a depolarization that inactivates Na+ conductance, and it supports those findings that assign this role to membrane shunting. Our results show that GABA can rapidly and selectively inhibit specific mechanoreceptors in the periphery. This type of peripheral inhibition may provide spiders with a mechanism for distinguishing between signals from potential prey, predators or mates, and responding with appropriate behaviour to each signal.

Predicting the responses of mechanoreceptor neurons to physiological inputs by nonlinear system identification.

AbstractThe nonlinear dynamic properties of action potential encoding were studied in mechanosensory neurons innervating the slits of a slit-sense organ in the tropical wandering spider, Cupiennius salei. The organ contains two types of neurons that are morphologically similar but have different dynamic properties. Type A neurons produce only one or two action potentials in response to a mechanical or electrical stimulus of any suprathreshold amplitude, while type B neurons can fire prolonged bursts of action potentials in response to similar stimuli. Neurons were stimulated with pseudorandomly modulated intracellular current while recording the resultant fluctuations in membrane potential and action potentials. A parallel cascade method was used to estimate a third-order Volterra series to describe the nonlinear dynamic relationship between membrane potential and action potentials. Kernels measured for the two types of neurons had reproducible forms that showed differences between the two neuron types. The measured kernels were able to predict the responses of the neurons to novel pseudorandomly modulated inputs with reasonable fidelity. However, the Volterra series did not adequately predict the difference in responses to step depolarizations.

Dendritic excitability and localization of GABA-mediated inhibition in spider mechanoreceptor neurons.

GABAergic inhibition of mechanosensory afferent axon terminals is a widespread phenomenon in vertebrates and invertebrates. Spider mechanoreceptor neurons receive efferent innervation on their peripherally located axons, somata and sensory dendrites, and the dendrites have recently been shown to be excitable. Excitability of the spider sensory neurons is inhibited by muscimol and GABA, agonists of ionotropic GABA receptors. Here we asked where in the neurons this inhibition occurs. We found no evidence for inhibition of action potentials in the sensory dendrites, but axonal action potentials were rapidly suppressed by both agonists. Earlier work showed that metabotropic GABAB receptors are located on the dendrites and distal somata of the spider sensory neurons, where they modulate voltage‐activated conductances and may provide slower, prolonged inhibition. Therefore, GABA released from single peripheral efferents may activate both ionotropic and metabotropic receptor types, providing rapid suppression of axonal activity followed by slower inhibition that eventually prevents action potential initiation in the distal dendrites.

Ca2+/calmodulin-dependent protein kinase II mediates the octopamine-induced increase in sensitivity in spider VS-3 mechanosensory neurons

G‐protein‐coupled octopamine (OA) receptors mediate their effects by Ca2+ signaling or adjusting intracellular cAMP levels. Depending on OA concentration and cell type, activation of OA receptors in excitable cells triggers excitatory or inhibitory effects, but the mechanisms by which Ca2+ or cAMP mediates these effects are not well understood. We investigated signaling mechanisms that are potentially activated by OA, and OA effects on excitability and frequency sensitivity in mechanosensory neurons innervating the VS‐3 slit sensilla on the patella of the spider Cupiennius salei. These neurons are directly innervated by octopaminergic efferents, and possess OA receptors that were immunoreactive to an antibody against an OA receptor highly expressed in mushroom bodies. OA application enhanced VS‐3 neuron sensitivity, especially at high stimulation frequencies. This enhancement lasted for at least 1 h after OA application. Changes in sensitivity were also detected when the Ca2+ ionophore ionomycin or the cAMP analog 8‐Br‐cAMP was applied. However, the cAMP pathway was unlikely to mediate the OA effect, as the protein kinase A inhibitor RP‐cAMPS did not diminish this effect. In contrast, the OA‐induced sensitivity enhancement was significantly reduced by KN‐62, an inhibitor of Ca2+/calmodulin‐dependent protein kinase II (CaMKII), and by the Ca2+ chelator BAPTA‐AM. OA depolarized the neurons by 3.8 mV from resting potential, well below the threshold for opening of voltage‐activated Ca2+ channels. OA also reduced the amplitudes of voltage‐activated K+ currents. We propose that OA receptors in VS‐3 neurons activate inositol 1,4,5‐trisphosphate, leading to Ca2+ release from intracellular stores. The Ca2+ surge switches on CaMKII, which modulates voltage‐activated K+ channels, resulting in persistent enhancement in excitability.

Random Stimulation of Spider Mechanosensory Neurons Reveals Long-Lasting Excitation by GABA and Muscimol

gamma-Aminobutyric acid type A (GABA(A)) receptor activation inhibits many primary afferent neurons by depolarization and increased membrane conductance. Deterministic (step and sinusoidal) functions are commonly used as stimuli to test such inhibition. We found that when the VS-3 mechanosensory neurons innervating the spider lyriform slit-sense organ were stimulated by randomly varying white-noise mechanical or electrical signals, their responses to GABA(A) receptor agonists were more complex than the inhibition observed during deterministic stimulation. Instead, there was rapid excitation, then brief inhibition, followed by long-lasting excitation. During the final excitatory phase, VS-3 neuron sensitivity to high-frequency signals increased selectively and their linear information capacity also increased. Using experimental and simulation approaches we found that the excitatory effect could also be achieved by depolarizing the neurons without GABA application and that excitation could override the inhibitory effect produced by increased membrane conductance (shunting). When the VS-3 neurons were exposed to bumetanide, an antagonist of the Cl(-) transporter NKCC1, the GABA-induced depolarization decreased without any change in firing rate, suggesting that the effects of GABA can be maintained for a long time without additional Cl(-) influx. Our results show that the VS-3 neurons response to GABA depends profoundly on the type of signals the neuron is conveying while the transmitter binds to its receptors.

Dynamic properties of Drosophila olfactory electroantennograms

Time-dependent properties of chemical signals are probably crucially important to many animals, but little is known about the dynamics of chemoreceptors. Behavioral evidence of dynamic sensitivity includes the control of moth flight by pheromone plume structure, and the ability of some blood-sucking insects to detect varying concentrations of carbon dioxide, possibly matched to host breathing rates. Measurement of chemoreceptor dynamics has been limited by the technical challenge of producing controlled, accurate modulation of olfactory and gustatory chemical concentrations over suitably wide ranges of amplitude and frequency. We used a new servo-controlled laminar flow system, combined with photoionization detection of surrogate tracer gas, to characterize electroantennograms (EAG) of Drosophila antennae during stimulation with fruit odorants or aggregation pheromone in air. Frequency response functions and coherence functions measured over a bandwidth of 0–100 Hz were well characterized by first-order low-pass linear filter functions. Filter time constant varied over almost a tenfold range, and was characteristic for each odorant, indicating that several dynamically different chemotransduction mechanisms are present. Pheromone response was delayed relative to fruit odors. Amplitude of response, and consequently signal-to-noise ratio, also varied consistently with different compounds. Accurate dynamic characterization promises to provide important new information about chemotransduction and odorant-stimulated behavior.

Shunting versus inactivation: Simulation of GABAergic inhibition in spider mechanoreceptors suggests that either is sufficient

Afferent neurons entering the central nervous systems of vertebrates and invertebrates receive presynaptic inhibition on their axon terminals. This usually involves an increase in membrane conductance (shunting) and depolarization (primary afferent depolarization, PAD). In arachnids and crustaceans the peripherally located parts of afferent neurons also receive efferent synapses. GABA (gamma-aminobutyric acid) plays a major role in both central and peripheral inhibition, activating chloride channels that depolarize the membrane and increase its conductance. Although both central and peripheral inhibition have been widely investigated, debate continues about the mechanisms involved, especially concerning the relative contributions of shunting versus inactivation of sodium channels by depolarization. Sensory neurons innervating spider VS-3 slit sensilla are accessible to intracellular recordings during mechanical or electrical stimulation. These neurons are inhibited by GABA, and both the electrophysiology and pharmacology of this inhibition have been studied previously. Here, we developed a Hodgkin-Huxley style model to simulate VS-3 neuron activity before and after GABA treatment. The model indicates that GABA-activated chloride current can entirely account for action potential suppression, and that either shunting or inactivation are sufficient to produce inhibition. This model also demonstrates that slowing of sodium current contributes to inhibition.