Biomedical subjects

L L Stockbridge

Publications and source records attributed to L L Stockbridge.

13 recordsLinked to original sources

PubMed1993

The morphology of the sensory neuron in the cockroach femoral tactile spine.

1. The cockroach femoral tactile spine contains a single bipolar sensory neuron. The mechanosensitive dendrite in the wall of the spine leads through the spine lumen to a cell body, and then to an axon that proceeds proximally along the femur. The ultrastructure of the sensory ending has been examined before with electron microscopy. However, the morphology of the complete neuron and its relationship to the general spine structure have not been described before. 2. The tactile spine neuron has been extensively used in electrophysiological studies, including intracellular recordings. Action-potential amplitudes and thresholds were variable and inversely related in intracellular recordings, which could be caused by variability in the location of the action-potential initiation region, the position of the recording electrode, or the neuronal morphology. Attempts to observe the complete neuronal morphology by dye injection were hampered by the opaque and autofluorescent cuticle surrounding the neuron. 3. We examined 10 tactile spine neurons, and their surrounding structures, by taking serial 1-micron sections through the base of the spine, normal to its long axis. The sections were examined with light microscopy, digitized by tracing onto a graphics tablet, and then reassembled with the use of computer software. Reconstructions were made of the borders of the spine cuticle, neuron, neuronal nucleus, glial wrappings, and the main trachea in the spine lumen. 4. There was considerable variability in the size and shape of the neuronal cell body, although the sensory dendrite and axon had more consistent morphologies.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

Subconductance states in calcium-activated potassium channels from canine airway smooth muscle.

The single-channel patch clamp technique was used to analyze subconductance states in the 260 pS calcium-activated potassium channel from canine airway smooth muscle. More than sixty minutes of single channel data (greater than 87,000 events) from five excised patches were analyzed. Six subconductance amplitudes were clearly established to be 17, 33, 41, 52, 63 and 72% of the full conductance. Subconductance openings were usually brief (milliseconds) and represented less than 5% of the total channel open time, but they also persisted for several seconds on rare occasions. They appeared to be unaffected by voltage or time after seal formation, but may have increased in occurrence with decreasing calcium concentration. Irregular amplitude intervals, and the presence of ramp-like, analog transitions between conductance states, suggest a model for maxi-K subconductance states in which the channel protein undergoes random conformational changes causing a variable pore size.

Animals↗

PubMed1991

The morphological basis of intracellular measurements in the cockroach tactile spine neuron.

The femoral tactile spine of the cockroach (Periplaneta americana) contains a single sensory neuron, which adapts rapidly and completely to step deformations of the spine. Techniques for stable intracellular recording from the tactile spine neuron have recently been established, allowing electrophysiological investigation of mechanotransduction and adaptation in this sensory neuron. However, intracellular recordings from the neuron produce a wide range of action potential heights and thresholds, raising the possibility that some penetrations are in adjacent, but closely coupled supporting glial cells. This problem is exacerbated because the cell cannot be visualized during penetration. Systematic measurements of action potential heights and thresholds were made in tactile spine cells, together with identification of some penetrated cells by intracellular injection of Lucifer Yellow. All stained cells were clearly sensory neurons, although their action potential amplitudes varied from 9 mV to 80 mV. Smaller action potentials were broader than larger action potentials, and the changes in height and shape could be explained by a simple cable conduction model using measured morphological and electrical parameters. The model could also account for the observed relationship between action potential height and threshold. These results indicate that reliable recording from the tactile spine neuron is possible, but that variability in the positions of the penetration or the spike initiating zone cause an apparently wide range of electrophysiological measurements.

Action Potentials↗

PubMed1991

Intracellular nonlinear frequency response measurements in the cockroach tactile spine neuron.

The threshold of the cockroach tactile neuron increases strongly with depolarization by a process involving at least two time constants. This effect is probably responsible for the rapid and complete adaptation of the neuron's response to step inputs. A technique for intracellular recording and stimulation of the neuron has recently been established and this allows direct observation of the dynamic response of the neuronal encoder. A white noise stimulus was used to modulate the membrane potential of the neuron. The first-order frequency response function between membrane potential and action potential discharge could be explained by a variable threshold model with two time constants. Second-order frequency response functions could be accounted for by a Wiener cascade model. The dynamic nonlinear behavior of the encoder can therefore be explained by a unidirectional threshold which increases linearly and dynamically with membrane potential.

Acoustic Stimulation↗

PubMed1990

Dissociation and culture of mechanosensory neurons for patch clamp analysis.

Single mechanosensory neurons were isolated from chordotonal organs of adult cockroach antennae. Transmission and scanning electron microscopy showed that the soma and part of the mechanosensory ending survived the dissociation. In culture, outgrowth occurred from the ending. Regions of cell membrane were accessible for patch clamp analysis and channels were recorded. The ability to record channel activity in isolated mechanosensory neurons will allow the study of mechanotransduction mechanisms at the membrane level.

Animals↗

PubMed1989

Characterization of a calcium-activated potassium channel in human fibroblasts.

The cell-attached and inside-out patch clamp techniques were used to record single-channel currents from human epidermal fibroblasts. A large-conductance channel (320 pS in symmetric 140 mM KCl) with high potassium selectivity was observed in many patches, particularly those located at the borders of the cells. The channel exhibited both voltage and calcium sensitivity and, therefore, was regarded as a variety of the large-conductance calcium-activated potassium channels reported in many preparations. Probability density functions, fitted to histograms of open and closed time durations at 35 degrees C, usually displayed a minimum of one open state and two closed states. However, kinetic analysis by the fractal method suggested more complicated behavior, particularly for the closed condition. It was not uncommon to observe several channels in one patch. This was distinguishable from the presence of subconductances, which were also observed. Although this channel could have many roles, it seems likely to mediate the calcium-activated conductance that underlies the hyperpolarizing response of fibroblasts to mechanical, electrical, or chemical stimuli.

Calcium↗

PubMed1988

Potassium channels in human and avian fibroblasts.

The cell-attached and excised inside-out patch-clamp techniques were used to study single-channel characteristics of potassium channels in cultured human and avian fibroblasts. Six different potassium channels were distinguished with conductances of 235 +/- 25, 190 +/- 57, 114 +/- 27, 77 +/- 14, 40 +/- 6 and 21 +/- 4 pS in symmetric 140 mM potassium solutions. The channels were separable by their conductances, ion-selectivities, voltage-sensitivities and kinetic properties. All six channels were found in both fully differentiated human skin fibroblasts and primary cultures of 72 h chick sclerotome. The largest channel (235 pS) had a steep bimodal voltage dependence, being open only around the resting membrane potential. It was imperfectly selective for potassium, having a relative sodium:potassium permeability of 0.3. The 190 pS channel was very potassium-selective, had an S-shaped voltage sensitivity and was calcium-dependent. The two intermediate-size channels (114 and 77 pS) had open probabilities of less than 0.5 under all of the conditions we used. They were not completely selective for potassium and were not voltage-sensitive. The two smallest channels (40 and 21 pS) were not well characterized. They both had open probabilities of less than 0.2 and showed no evidence of voltage-sensitivity. The 40 pS channel seemed highly potassium-selective. A suction stimulus was used to test all observed channels for mechanosensitivity but none of the six potassium channels was mechanosensitive. Another small channel, with very clear mechanical sensitivity, was seen on a few occasions; this channel has not yet been characterized.

Animals↗

PubMed1988

Properties of synaptic transmission at the neuromuscular junction of the squid, Loligo opalescens.

1. Spontaneous and evoked synaptic activity were recorded from the muscles of squid fin and mantle. These spontaneous synaptic potentials were large (up to 30 mV) and pleomorphic. Their amplitudes were not normally distributed, nor did they appear to be clustered in integral multiples of some "unit" event size. 2. Electrical stimulation of the nerve resulted in muscle twitches when the bath calcium concentration was a third normal or higher. The frequency of spontaneous synaptic events was unaffected by low calcium. 3. The large size of spontaneous events may mean that the synchronized release of only a few such "quanta" are sufficient to cause muscle action potentials and contraction. 4. The shapes of spontaneous events correlated poorly with their amplitudes, which is consistent with release from multiple synaptic sites with distinct properties.

Animals↗

PubMed1988

Stretch-activated cation channels in human fibroblasts.

Nonconfluent fibroblasts are relatively depolarized when compared with confluent fibroblasts, and transient hyperpolarizations result from a range of external stimuli as well as internal cellular activities. This electrical activity ceases, along with growth and mitogenic activity, when the cells become confluent. A calcium-activated potassium conductance is thought to be responsible for these hyperpolarizations, but in human fibroblasts the large calcium-activated potassium channel is not stretch-activated. We report here the identification of single stretch-activated cation channels in human fibroblasts, using the cell-attached and inside-out patch clamp techniques. The most prominent channel had a conductance of approximately 60 pS (picoSeimens) in 140 mM potassium and was permeable to potassium and sodium. The channel showed significant adaptation of activity when stretch was maintained over a period of several seconds, but a static component persisted for much longer periods. Higher conductance channels were also observed in a few excised patches.

Cations↗

PubMed1988

Fractal and Markov behavior in ion channel kinetics.

Kinetic analysis of ion channel recordings attempts to distinguish the number and lifetimes of channel molecular states. Most kinetic analysis assumes that the lifetime of each state is independent of previous channel history, so that open and closed durations are Markov processes whose probability densities are sums of exponential decays. An alternative approach assumes that channel molecules have many configurtions with widely varying lifetimes. Rates of opening and closing then vary with the time scale of observation, leading to fractal kinetics. We have examined kinetic behavior in two types of channels from human and avian fibroblasts, using a maximum likehood method to test the dependence of rates on observational time scale. For both channels, openings showed mixed fractal and Markov behavior, while closings gave mainly fractal kinetics.

Animals↗

PubMed1988

Differential conduction at axonal bifurcations. I. Effect of electrotonic length.

1. Frequency-dependent differential conduction of action potentials into one daughter branch of the squid giant axon is demonstrated. 2. This differential conduction arises from differences in the electrotonic length of the daughters, rather than inhomogeneities in membrane properties. 3. It may therefore be a more general phenomenon in the nervous system than is differential conduction which does depend upon membrane inhomogeneities.

Action Potentials↗

PubMed1986

A TTX-resistant propagating calcium action potential.

The cross-commissural (CC) cell in the supraesophageal ganglion of the giant barnacle, Balanus nubilus, was stimulated intrasomatically and antidromically in normal saline and 3 X 10(-7) M tetrodotoxin (TTX) saline. The action potential in normal saline contained both sodium and calcium components, each independently capable of propagation. Evidence that the action potential in TTX saline was calcium dependent included: the amplitude of the spike in TTX saline increased monotonically with increasing calcium; it was blocked by the calcium channel blockers La, Ni, Cd and Co; and equimolar substitution of Ba or Sr for Ca in TTX saline supported regenerative activity. Evidence that the calcium component could propagate alone included: the distance over which the calcium action potential traveled exceeded the space constant of the axon; the biphasic nature of the extracellularly recorded action potential, which propagated to the axon in the nerve root, indicated inward regenerative current was occurring on the axon; electrotonically spread potentials were clearly distinguishable from active regenerative potentials. In addition, optical experiments using the calcium indicator dye arsenazo III (28) showed that the relative magnitude of the calcium signal was not diminished along the axon in TTX saline compared with normal saline. The critical external calcium concentration necessary to support the propagating action potential in TTX saline was estimated to be between 1.25 and 5 mM. To our knowledge, this is the first direct observation of a neuron with sodium and calcium channels of apparently normal kinetics where the calcium component alone can propagate in the absence of an outward current blocker. Our results suggest that there is a greater density of calcium channels on the CC axon than on the axons of other neurons.

Animals↗

PubMed1986

Regional properties of calcium entry in barnacle neurons determined with Arsenazo III and a photodiode array.

Calcium changes were simultaneously measured at many positions on individual neurons from the supraesophageal ganglion of the barnacle by detecting absorbance changes of the indicator dye Arsenazo III with a 10 X 10 photodiode array. These changes were correlated with positions on the stimulated cell determined from Lucifer yellow injections. Absorbance signals were found at all locations on the cells, demonstrating that calcium channels were distributed on the somata, axons, and neuropil processes. By comparing the amplitude of the signal with the membrane area in each region, we could measure the calcium entry per impulse per unit of surface in each part of the cell. Assuming that the properties of the calcium channels are the same in all regions, we determined that calcium channels were distributed uniformly along the commissural axon of one cell and were found at higher density in the neuropil. Because significant calcium changes are only detected when cells are depolarized above about -20 mV, the presence of absorbance signals indicates membrane depolarization above this level. We used this fact to show that calcium spikes propagate along the axon and into the neuropil of one cell, along the axon of another, and not at all in a third. Differences in time course of calcium transients were observed in different regions of cells. The recovery time course was faster at the edge of the cell body than in the center and faster in the neuropil than in the axon or soma. During trains of action potentials or during wide action potentials in tetraethylammonium (TEA) saline, the calcium signal reached a plateau in the neuropil while continuing to rise in the axon and soma.

Action Potentials↗