Ulli Höger

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.

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.

Acetylcholine receptors in spider peripheral mechanosensilla

Peripherally located parts of spider mechanosensory neurons are modulated by several neurotransmitters released from apposed efferent fibers. Activities of acetylcholine (ACh) synthesizing enzyme choline acetyltransferase (ChAT) and ACh degrading enzyme acetylcholine esterase (AChE) were previously found in some efferent fibers. ChAT activity was also present in all the mechanosensory neurons, while AChE activity was only found in some. We show that spider mechanosensory neurons and probably some efferent neurons are immunoreactive to a monoclonal antibody against muscarinic ACh receptors (mAChRs). However, application of muscarinic agonists did not change the physiological responses or membrane potentials of neurons in the lyriform organ VS-3. Similarly, the sensitivities of the neurons of trichobothria (filiform hairs) remained unchanged after application of these agonists. Therefore, activation of mAChRs may only modulate the function of spider mechanosensory neurons indirectly, for example, by affecting the release of other transmitter(s). However, a subgroup of VS-3 neurons was inhibited by ACh, which also depolarized the membrane similar to these neurons’ responses to GABA, suggesting that ACh activates anion channels in these neurons. Interestingly, all of the neurons responding to ACh were the rapidly adapting Type A neurons that were previously shown to express AChE activity.

Calcium concentration changes during sensory transduction in spider mechanoreceptor neurons.

Most mechanoreceptor neurons encode mechanical signals into action potential trains within the same cell. Evidence suggests that intracellular calcium ion concentration, [Ca2+], increases during mechanotransduction, either by direct entry through mechanically activated channels or indirectly through voltage‐activated calcium channels. However, little is known about the amounts of calcium involved or its roles in mechanotransduction. We estimated [Ca2+] in mechanoreceptor neurons of the spider, Cupiennius salei, during mechanical stimulation using Oregon Green BAPTA‐1, and a single‐compartment model of [Ca2+] as a function of action potential firing rate. Resting [Ca2+] was approximately 400 nm and increased to up to 2 µm at 30 action potentials/s. Similar levels of resting and stimulated [Ca2+] were obtained in the cell soma, axon and two parts of the sensory dendrite, including the region immediately adjacent to the site of sensory transduction. The time constant of rise and fall of [Ca2+] was 1–5 s in the dendrite and axon, but up to 15 s in the soma. Calcium elevation was dependent on action potentials and could not be induced by the receptor potential alone. Blockade of voltage‐activated calcium channels by nickel ions prevented calcium increase, but thapsigargin, which empties intracellular calcium stores, had no effect. Estimates of calcium entry per action potential from fluorescence changes agreed approximately with estimates based on action potential voltage–time profile and previous reports of calcium channel properties. This first report of calcium levels during transduction in spiking mechanoreceptors suggests that calcium signaling plays important roles in primary somatosensory neurons.

Extracellular acid increases the open probability of transduction channels in spider mechanoreceptors

Ion channels of the epithelial sodium channel, degenerin and acid‐sensitive channel (ENaC/DEG/ASIC) family share a number of structural and functional homologies. Several members of this group have been linked to mechanoreception and nociception, but there is no direct evidence that these molecules cause the transduction of mechanical stimuli in any mechanoreceptor. The receptor channels of a spider mechanoreceptor, the VS‐3 slit‐sense organ of Cupiennius salei, show several similarities to ENaC/DEG/ASIC channels, including Na+ selectivity and amiloride blockade. We recorded the receptor current under voltage clamp in VS‐3 neurons at different extracellular pH values. Acid pH partially blocked the delayed rectifier K+ current and increased the receptor current in these cells. Noise analysis of the receptor current showed that low pH increased the open probability of the receptor channels. Therefore, acid sensitivity is a further similarity between these mechanoreceptor channels and the ENaC/DEG/ASIC family.

Estimated single-channel conductance of mechanically-activated channels in a spider mechanoreceptor

Noise analysis was used to estimate the single-channel conductance and number of channels responsible for the mechanically-activated current in the sensory neurons of a spider mechanoreceptor organ. External slits of the VS-3 slit-sense organ in the patellar cuticle of Cupiennius salei were moved with a piezoelectric stimulator while glass microelectrodes penetrated the adjacent cell bodies. Receptor currents were measured by the switching single-electrode voltage clamp technique during both step and ramp displacements of the slits. Current records were segmented in time, and the variance and amplitude of the current were obtained from each segment, to allow fitting of the variance vs. amplitude relationship by a standard equation based on a two-state channel. Mean values of 7.5 pS and 253 were obtained for the conductance and number of channels from 75 separate recordings. These values are in good agreement with the small number of other estimates of these parameters from different mechanoreceptor preparations.

Ratiometric calcium concentration estimation using LED excitation during mechanotransduction in single sensory neurons

In a previous study using Oregon Green BAPTA-1 fluorescence we found that intracellular calcium concentration in spider mechanoreceptor neurons rose during mechanical stimulation. We also showed that calcium elevation required the opening of voltage-dependent calcium channels by action potentials, and could not be produced by the receptor potential alone. While evidence for mechanisms of calcium elevation in these neurons was clear, our estimates of actual calcium concentration depended on properties of the fluorescent dye in the neuron cytoplasm that could not be verified. We have now developed a method for ratiometric estimation of calcium concentration in these neurons using Fura Red dye, excitation by two light emitting diodes (LEDs) of different wavelengths, and an avalanche photodiode fluorescence detector. The method is simple and economical to implement, allows concentration changes to be measured in the millisecond time range, and could easily be applied to a wide range of preparations. Resting calcium concentration in these neurons was about 70nM and rose to a maximum of about 400nM at firing rates above 20 action potentials per second.

Principal dynamic mode analysis of action potential firing in a spider mechanoreceptor

The encoding of mechanical stimuli into action potentials in two types of spider mechanoreceptor neurons is modeled by use of the principal dynamic modes (PDM) methodology. The PDM model is equivalent to the general Wiener–Bose model and consists of a minimum set of linear dynamic filters (PDMs), followed by a multivariate static nonlinearity and a threshold function. The PDMs are obtained by performing eigen-decomposition of a matrix constructed using the first-order and second-order Volterra kernels of the system, which are estimated by means of the Laguerre expansion technique, utilizing measurements of pseudorandom mechanical stimulation (input signal) and the resulting action potentials (output signal). The static nonlinearity, which can be viewed as a measure of the probability of action potential firing as a function of the PDM output values, is computed as the locus of points of the latter that correspond to output action potentials. The performance of the model is assessed by computing receiver operating characteristic (ROC) curves, akin to the ones used in decision theory and quantified by computing the area under the ROC curve. Three PDMs are revealed by the analysis. The first PDM exhibits a high-pass characteristic, illustrating the importance of the velocity of slit displacement in the generation of action potentials at the mechanoreceptor output, while the second and third PDMs exhibit band-pass and low-pass characteristics, respectively. The corresponding three-input nonlinearity exhibits asymmetric behavior with respect to its arguments, suggesting directional dependence of the mechanoreceptor response on the mechanical stimulation and the PDM outputs, in agreement to our findings from a previous study (Ann Biomed Eng 27:391–402, 1999). Differences between the Type A and B neurons are observed in the zeroth-order Volterra kernels (related to the average firing), as well as in the magnitudes of the second and third PDMs that perform band-pass and low-pass processing of the input signal, respectively.

Feedback modulation of transduction by calcium in a spider mechanoreceptor

Calcium ions play important roles in the adaptation of auditory hair cells, and there is evidence that they are involved in modifying the sensitivity and adaptation of a variety of vertebrate and invertebrate mechanoreceptors. However, there is little direct evidence concerning the concentration changes, signaling pathways or ultimate effects of these proposed modulatory mechanisms. We measured receptor potential, receptor current and action potentials intracellularly during mechanotransduction in a group of sensory neurons of the spider Cupiennius salei, which possesses low‐voltage‐activated calcium channels. Simultaneously, we elevated intracellular [Ca2+] by UV light release from cage molecules, and observed increases in [Ca2+] as changes in calcium‐sensitive dye fluorescence. Increases of 10–15% in [Ca2+] caused reductions of approximately 40% in receptor potential and approximately 20% in receptor current. Mechanically evoked action potential firing caused much larger increases in [Ca2+], and the firing rate fell as [Ca2+] rose during mechanical stimulation. Release of caged calcium just before mechanical stimulation significantly reduced peak firing. Dose–response measurements suggested that the binding of one or two intracellular calcium ions per channel reduces the probability of the mechanotransduction channel being open. Our data indicate that calcium regulates sensitivity in these mechanoreceptor neurons by negative feedback from action potentials onto transduction channels.

Regional distribution of calcium elevation during sensory transduction in spider mechanoreceptor neurons.

Spider mechanosensory VS-3 neurons receive peripheral efferent synaptic modulation, with regional variations in the types of efferent synapses and transmitter receptors. VS-3 somata possess a voltage-activated calcium current, but the levels and time courses of calcium changes in other regions are unknown. The roles of calcium in these neurons are not completely understood, but could include modulation of both mechanosensitivity and response dynamics. Here, we measured calcium concentration rises caused by single, mechanically induced action potentials in VS-3 sensory dendrites, somata and axons, using Oregon Green BAPTA-1 fluorescence. Calcium concentration rose by approximately 1 nM following each action potential. Time courses of calcium rise and fall were similar in the three regions but the rise in amplitude was about 50% higher in the sensory dendrite than in the soma. Antibody to the Ca(V)3.1(alpha(1g)) isotype of T-type calcium channel labeled all three neuronal regions. Some Ca(V)3.1 labeling colocalized with synapsin labeling, suggesting that calcium channels play some part in efferent modulation. We conclude that mechanically stimulated action potentials start near sensory dendrite tips and pass rapidly through the neurons to the axons, activating low voltage activated calcium channels in all three regions and causing calcium concentration to rise rapidly in each region. These results suggest important roles for calcium in several stages of mechanosensation.