A neuromuscular transmission block produced by a cancer tissue extract derived from a patient with the myasthenic syndrome.

The myasthenic syndrome occasionally is associated with bronchogenic carcinoma. The neuromuscular transmission defect in this syndrome is characterized by a reduction of acetylcholine release from motor nerve endings. This paper reports that an acetone extract of cancer tissue from a patient with the syndrome reduces the acetylcholine release from motor nerve endings and produces a neuromuscular transmission defect in the frog nerve-muscle preparation. This suggests that the pathogenic substance(s) contained in the extract may be produced by certain types of bronchogenic carcinoma and may cause the myasthenic syndrome.

Changes in the axonal conduction velocity of pyramidal tract neurons in the aged cat.

The present study was undertaken to determine whether age-dependent changes in axonal conduction velocity occur in pyramidal tract neurons. A total of 260 and 254 pyramidal tract neurons were recorded extracellularly in the motor cortex of adult control and aged cats, respectively. These cells were activated antidromically by electrical stimulation of the medullary pyramidal tract. Fast- and slow-conducting neurons were identified according to their axonal conduction velocity in both control and aged cats. While 51% of pyramidal tract neurons recorded in the control cats were fast conducting (conduction velocity greater than 20 m/s), only 26% of pyramidal tract neurons in the aged cats were fast conducting. There was a 43% decrease in the median conduction velocity for the entire population of pyramidal tract neurons in aged cats when compared with that of pyramidal tract neurons in the control cats (P < 0.001, Mann-Whitney U-test). A linear relationship between the spike duration of pyramidal tract neurons and their antidromic latency was present in both control and aged cats. However, the regression slope was significantly reduced in aged cats. This reduction was due to the appearance of a group of pyramidal tract neurons with relatively shorter spike durations but slower axonal conduction velocities in the aged cat. Sample intracellular data confirmed the above results. These observations form the basis for the following conclusions: (i) there is a decrease in median conduction velocity of pyramidal tract neurons in aged cats; (ii) the reduction in the axonal conduction velocity of pyramidal tract neurons in aged cats is due, in part, to fibers that previously belonged to the fast-conducting group and now conduct at slower velocity.

An alternative method for the analysis of neuron passive electrical data which uses integrals of voltage transients.

The traditional method for analyzing passive electrical data from neurons when specific morphological data are unavailable consists of decomposing the voltage response of the cell into a series of exponential functions (the peeling method) and substituting the time constants of these exponential functions into equations derived from cable theory (Rall W, Core conductor theory and cable properties of neurons. In: Handbook of Physiology. The Nervous System. Cellular Biology of Neurons. Bethesda, MD. Am Physiol Soc. Section 1, Part 1, 1977;1(3):39-97). In the present report, an alternative method is examined for analyzing these kinds of data, the integrals of transients method (Eisenberg RS, Mathias RT. Structural analysis of electrical properties of cells and tissues. CRC Critical Reviews in Bioengineering 1980;4:203-232). The integrals required are easily obtained from input resistance data and any theoretical model that is appropriate for the neurons under study can be used, provided that the impedance function can be determined. In order to demonstrate this alternative method, a simple 3-compartment model with both dendritic taper and somatic shunt is used to model data obtained from fast-type alpha-motoneurons in the spinal cord of the cat. These results are compared with results obtained using the traditional peeling method. This comparison indicates that passive electrical data from fast-type motoneurons are best analyzed using a theoretical model that includes both dendritic taper and somatic shunt. Furthermore, our results show that the integrals of transients method can facilitate this analysis.

Neuronal network analysis based on arrival times of active-sleep specific inhibitory postsynaptic potentials in spinal cord motoneurons of the cat.

The neuronal network responsible for motoneuron inhibition and loss of muscle tone during active (REM) sleep can be activated by the injection of the cholinergic agonist carbachol into a circumscribed region of the brainstem reticular formation. In the present report, we studied the arrival times of inhibitory postsynaptic potentials (IPSPs) observed in intracellular recordings from cat spinal cord motoneurons. These recordings were obtained during episodes of motor inhibition induced by carbachol or during motor inhibition associated with naturally occurring active sleep. When the observed IPSP arrival times were analyzed as a superposition of renewal processes occurring in a pool of pre-motor inhibitory interneurons, it was possible to estimate the following parameters: (1) the number of independent sources of the IPSPs; (2) the rate at which each source was bombarded with excitatory postsynaptic potentials (EPSPs); and (3) the number of EPSPs required to bring each source to threshold. From the data based upon the preceding parameters and the unusually large amplitudes of the active sleep-specific IPSPs, we suggest that each source is a cluster of synchronously discharging pre-motor inhibitory interneurons. The analysis of IPSP arrival times as a superposition of renewal processes, therefore, provides quantitative information regarding neuronal activity that is as far as two synapses upstream from the site of the recording electrode. Consequently, we suggest that a study of the temporal evolution of these parameters could provide a basis for dynamic analyses of this neuronal network and, in the future, for other neuronal networks as well.

Neurotrophic effects on the electrical properties of cultured muscle produced by conditioned medium from spinal cord explants.

The passive electrical properties of cultured chick skeletal muscle are significantly altered when the muscle is co-cultured with spinal cord explants. A reduced transverse membrane resistance appears to be responsible for the smaller values of input resistance, space constant, and time constant observed in co-cultures relative to those observed in pure muscle cultures. In this report, we show that neuromuscular junctions are not required in order to observe this neurotrophic effect because medium from spinal cord explant cultures is capable of producing the same reduction in transverse membrane resistance as the co-culturing of spinal cord explants with muscle. Control medium from liver explant cultures has no effect on muscle passive electrical properties. These results indicate that a trophic substance which is capable of regulating the electrical properties of excitable cells is released into the culture medium by spinal cord explants.

Neuronal-like features of TE671 cells: presence of a functional nicotinic cholinergic receptor.

The human medulloblastoma cell line TE671 has been investigated and found to have several 'neuron-like' properties, including the presence of a functional nicotinic receptor. The cell line TE671 is composed of at least 5 stable morphologic cell types. Resting potentials recorded with intracellular microelectrodes were low (-17 mV to -31 mV) but all cell types were capable of generating Na+-dependent action potentials following anode-brake stimulation. Rare spontaneous hyperpolarizing potentials, suggestive of synaptic activity, were also observed. TE671 cells were completely unresponsive to iontophoresed GABA but did respond to acetylcholine (ACh). The most common response to ACh was a depolarization accompanied by an increase in membrane conductance. When large amounts of ACh were delivered, depolarization followed by hyperpolarization was frequently observed. Depolarizing responses to ACh are abolished in Na+-free solution while hyperpolarizing responses to ACh were still present following the removal of both Na+ and Cl- from the bathing solution. The depolarization to ACh is mediated through a nicotinic cholinergic receptor. Depolarization was completely blocked in the presence of 10(-6) M alpha-bungarotoxin, 4.4 x 10(-5) M D-tubocurarine, or 10(-4) M decamethonium. Atropine was only 50% effective at 10(-4) M and hexamethonium was ineffective at 10(-4) M. In vitro binding of receptor ligands to membranes prepared from TE671 cells revealed high levels of [125I]alpha-bungarotoxin (alpha-BuTx) binding sites, in addition to lower levels of other ligand binding sites. [125I]alpha-BuTx bound to a single, saturable high affinity site in either membrane preparations or intact TE671 cells. Binding was potently inhibited by the classical nicotinic acetylcholine receptor antagonists D-tubocurarine and decamethonium. Nicotine and carbamylcholine showed intermediate potencies in inhibiting binding while atropine and hexamethonium showed little ability to inhibit [125I]alpha-BuTx binding. The data obtained from [125I]alpha-BuTx binding studies agree qualitatively with the electrophysiological data on the depolarizing ACh response and together they provide strong evidence that TE671 cells possess a functional nicotinic acetylcholine receptor. This cell may therefore be useful as a stable source with which to characterize mammalian neuronal nicotinic acetylcholine receptors and membrane events related to its activation.

Passive electrical properties of motoneurons in aged cats following axotomy.

The objective of this study was to determine whether the aging process influences the changes in the electrophysiological properties of motoneurons that occur as a consequence of axotomy. Accordingly, using intracellular recording and stimulating techniques, the basic electrical properties of control (unaxotomized) and axotomized spinal cord motoneurons of aged cats were determined. Compared with control motoneurons, axotomized motoneurons exhibited increases in input resistance (Rin), membrane time constant (tau b) and the equalizing time constant (tau c). While the electrotonic length (L) remained unchanged, axotomy induced a decrease in the total cell capacitance (Ccell). The post-axotomy reduction of Ccell indicates that the motoneuron surface area was reduced and the increased membrane time constant indicates that there was an increase in membrane resistivity (Rm). The post-axotomy conservation of L accompanied by an increase in Rm suggests that aged axotomized motoneurons undergo geometrical changes. Furthermore, calculations based on cable theory suggest that the diameter of the equivalent cylinder (d) decreased following axotomy, whereas the equivalent cylinder length (l) remained unaffected. It is concluded that axotomy produces significant alterations in the soma-dendritic portion of aged spinal motoneurons, as indicated by the changes found in their passive electrophysiological properties, and that the pattern of the response that occurs in axotomized motoneurons of adult cats is also present in axotomized motoneurons of aged animals.

Experimental analysis of the method of 'peeling' exponentials for measuring passive electrical properties of mammalian motoneurons.

Trigeminal motoneurons of the guinea pig brain stem slice preparation were studied using intracellular recording techniques. The voltage response to a 100-ms constant-current pulse was studied and a population of cells was found which did not exhibit sag or overshoot of their voltage response to a pulse of hyperpolarizing current of < 1 nA but did exhibit both phenomena when a current pulse of > 1 nA was used. The sag and overshoot observed with large-current pulses were reduced or blocked when 4 mM CsCl was added to the bathing solution. This observation supports the hypothesis that these phenomena were due to the voltage- and time-dependent activation of the Q-current. The method of peeling exponentials was then used to correct raw voltage data from cells in which the Q-current was present. The mean membrane time constant was within 1% and the mean input resistance was within 2% of the means for these parameters when measured in these same cells under conditions in which the Q-current was absent. We conclude from these experiments that the method of peeling exponentials is valid for obtaining estimates of the membrane time constant and input resistance from cells that exhibit sag and overshoot due to voltage- and time-dependent changes in the magnitude of the Q-current.

Age-dependent changes in cat masseter nerve: an electrophysiological and morphological study.

The present study was undertaken to determine the manner in which aging affects the function and structure of the masseter nerve in old cats. Electrophysiological data demonstrated a significant decrease in the conduction velocity of the action potential in old cats compared with that observed in adult cats. Light microscopic analyses revealed an age-dependent decrease in axon diameter. Electron microscopic observations of the masseter nerve in the aged cats revealed a disruption of the myelin sheaths and a pronounced increase in collagen fibers in the endoneurium and perineurium. These morphological changes are discussed and then related to the decrease in conduction velocity which was observed in the electrophysiological portion of this study.

Cell size and geometry of spinal cord motoneurons in the adult cat following the intramuscular injection of adriamycin: comparison with data from aged cats.

Adriamycin (ADM), an antineoplastic antibiotic, when injected intramuscularly, is taken up by motoneuron axonal terminals and retrogradely transported to the motoneuron soma where it exerts its neurotoxic effect. In the present study, ADM was injected into the hindlimb muscles of five adult cats. Measurements of the electrophysiological properties of the lumbar motoneurons innervating these muscles were obtained using intracellular techniques. Based upon these data the equivalent cylinder model of motoneurons was employed to evaluate ADM-induced changes in cell size and cell geometry. The size of cell somas in the ventral horn was also measured using light microscopy and computer imaging software. There were significant increases in the membrane time constant (25%) and input resistance (50%) in motoneurons whose muscles were treated with ADM (ADM-MNs) compared with data from control motoneurons (control-MNs). The increase in membrane time constant is attributed to an increase in membrane resistance; the increase in input resistance appears to depend upon both an increase in membrane resistance and a decrease in total cell surface area. Cell capacitance, which is proportional to the total cell surface area, was significantly reduced (15%) in ADM-MNs. Calculations based on cable theory indicate that while there was no significant change in the length of the equivalent cylinder for ADM-MNs, there was a significant decrease (17%) in the diameter of the equivalent cylinder. These data indicate that there is a decrease in total cell surface area which can be attributed to the shrinkage of branches throughout the dendritic tree. There was also a small (7%) but statistically significant decrease in the electrotonic length of ADM-MNs. Morphological analysis also revealed that the mean cross-sectional area of the somas of those ventral horn neurons which are likely to correspond to the motoneuron population was significantly reduced on the ADM-treated side compared to that of neurons on the control side. We conclude that significant geometrical changes were induced in lumbar motoneurons of adult cats after ADM was injected to their muscles. In old cats, spinal cord motoneurons exhibit similar patterns of changes in their electrophysiological characteristics which have also been suggested to be correlated with changes in cell geometry. The question then arises as to whether the response of motoneurons to ADM and the aging process reflects a stereotypic reaction of motoneurons to a variety of insults or whether the response to ADM mirrors specific aspects of the aging process.

Changes in electrophysiological properties of cat hypoglossal motoneurons during carbachol-induced motor inhibition.

The control of hypoglossal motoneurons during sleep is important from a basic science perspective as well as to understand the bases for pharyngeal occlusion which results in the obstructive sleep apnea syndrome. In the present work, we used intracellular recording techniques to determine changes in membrane properties in adult cats in which atonia was produced by the injection of carbachol into the pontine tegmentum (AS-carbachol). During AS-carbachol, 86% of the recorded hypoglossal motoneurons were found to be postsynaptically inhibited on the basis of analyses of their electrical properties; the electrical properties of the remaining 14% were similar to motoneurons recorded during control conditions. Those cells that exhibited changes in their electrical properties during AS-carbachol also displayed large-amplitude inhibitory synaptic potentials. Following sciatic nerve stimulation, hypoglossal motoneurons which responded with a depolarizing potential during control conditions exhibited a hyperpolarizing potential during AS-carbachol. Both spontaneous and evoked inhibitory potentials recorded during AS-carbachol were comparable to those that have been previously observed in trigeminal and spinal cord motoneurons under similar experimental conditions as well as during naturally occurring active sleep. Calculations based on modeling the changes that we found in input resistance and membrane time constant with a three-compartment neuron model suggest that shunts are present in all three compartments of the hypoglossal motoneuron model. Taken together, these data indicate that postsynaptic inhibitory drives are widely distributed on the soma-dendritic tree of hypoglossal motoneurons during AS-carbachol. These postsynaptic inhibitory actions are likely to be involved in the pathophysiology of obstructive sleep apnea.

Passive electrophysiological properties of aged and axotomized cat spinal cord motoneurons: the effect of cell size and electrode shunt.

Intracellular recordings were obtained from intact and axotomized lumbar motoneurons of aged cats. The sub-threshold electrical properties of these cells were measured, including input resistance, resting membrane potential, and the first two equalizing time constants as well as their associated amplitude constants. These data were used in a semi-infinite cable model of the motoneuron to estimate the size of the shunt resistance (Rshunt) which is created when the electrode penetrates the cell membrane. The average Rshunt for intact aged cells was 5.35 +/- 1.01 M omega, while that for the axotomized aged cells was 8.93 +/- 1.20 M omega. The statistically significant difference in mean shunt magnitude did not affect the measurements of membrane time constant because this constant is independent of the shunt in this model of the motoneuron. However, the determination of cell input resistance, which is not independent of the shunt, was shown to underestimate the real cell input resistance by 23-29%. We therefore conclude that the shunt resistance is an important factor which should be taken into account when measuring input resistance.

Age-related changes in soma size of neurons in the spinal cord motor column of the cat.

The present study was undertaken to examine the effect of the aging process on the soma size and number of motoneurons and interneurons in the motor column of the spinal cord of old cats. Neurons in the motor column were divided into small and large populations based on a bimodal distribution of their soma cross-sectional areas. A 17% decrease in the cross-sectional area of small neurons was observed, this decrease was statistically significant (P < 0.0001). The cross-sectional area of large neurons decreased by only 6%, which was statistically significant (P < 0.05). On the other hand, there was no significant difference in the number of large, small or of these combined population of ventral horn neurons in the aged cats compared with the control animals. This data suggest that neurons in the motor column are not uniformly affected by the aging process because morphological changes are proportionally greater in small neurons than in large neurons.

Renshaw cells are inactive during motor inhibition elicited by the pontine microinjection of carbachol.

The present study was undertaken to determine whether the postsynaptic inhibition of motoneurons that occurs following the pontine microinjection of carbachol in the decerebrate cat is due to the activity of Renshaw cells. Thirty-two out of 37 Renshaw cells (86%) were spontaneously active prior to the administration of carbachol, whereas only 2 out of 13 Renshaw cells (15%) discharged during carbachol-induced motor inhibition. In addition, discrete inhibitory synaptic potentials were observed in 33% of the Renshaw cells from which intracellular recordings were obtained after carbachol administration, indicating that these cells were actively inhibited. The finding that a population of Renshaw cells, which inhibit motoneurons, were themselves inhibited during a period of profound motoneuron inhibition was quite unexpected. These results support the conclusion that Renshaw cells are not the inhibitory interneurons that are responsible for the powerful inhibition of motoneurons that occurs following the pontine microinjection of carbachol.

Cable properties of spinal cord motoneurons in adult and aged cats.

1. The electrophysiological properties of spinal cord alpha-motoneurons were investigated in adult cats (1-3 yr old) and old cats (14-15 yr old) using intracellular recording techniques. Voltage transients following depolarizing pulses of current were analyzed according to the procedure described by Ito and Oshima (15). The input resistance of each cell, together with the passive electrical time constants, were used to estimate the electrotonic length and total cell capacitance of each motoneuron. 2. Adult and old motoneurons both exhibited an undershoot of the membrane potential following the cessation of a subthreshold depolarizing current pulse (15). The average time constant for the decay of this undershoot in membrane potential was statistically indistinguishable in motoneurons of adult and aged animals. 3. The average membrane time constant of motoneurons in aged cats was 19% longer than that of motoneurons in adult cats. 4. The average total cell capacitance of motoneurons in aged cats was 16% smaller than that of motoneurons in adult cats. 5. The average electrotonic length of old motoneurons was statistically indistinguishable from that of motoneurons in adult cats. 6. From these results, we conclude that there is an age-dependent increase in the membrane resistance and an age-dependent decrease in cell surface area of alpha-motoneurons of the lumbar spinal cord in aged cats. Both of these phenomena are believed to contribute to the age-dependent increase in input resistance that has been previously reported to occur in motoneurons in aged cats (18).

Motoneuron properties during motor inhibition produced by microinjection of carbachol into the pontine reticular formation of the decerebrate cat.

It is well established that cholinergic agonists, when injected into the pontine reticular formation in cats, produce a generalized suppression of motor activity (1, 3, 6, 14, 18, 27, 33, 50). The responsible neuronal mechanisms were explored by measuring ventral root activity, the amplitude of the Ia-monosynaptic reflex, and the basic electrophysiological properties of hindlimb motoneurons before and after carbachol was microinjected into the pontine reticular formation of decerebrate cats. Intrapontine microinjections of carbachol (0.25-1.0 microliter, 16 mg/ml) resulted in the tonic suppression of ventral root activity and a decrease in the amplitude of the Ia-monosynaptic reflex. An analysis of intracellular records from lumbar motoneurons during the suppression of motor activity induced by carbachol revealed a considerable decrease in input resistance and membrane time constant as well as a reduction in motoneuron excitability, as evidenced by a nearly twofold increase in rheobase. Discrete inhibitory postsynaptic potentials were also observed following carbachol administration. The changes in motoneuron properties (rheobase, input resistance, and membrane time constant), as well as the development of discrete inhibitory postsynaptic potentials, indicate that spinal cord motoneurons were postsynaptically inhibited following the pontine administration of carbachol. In addition, the inhibitory processes that arose after carbachol administration in the decerebrate cat were remarkably similar to those that are present during active sleep in the chronic cat. These findings suggest that the microinjection of carbachol into the pontine reticular formation activates the same brain stem-spinal cord system that is responsible for the postsynaptic inhibition of alpha-motoneurons that occurs during active sleep.