Functional coupling between motor and sensory nerves through contraction of sphincters in the pudendal area of the female cat.

The question of whether skin receptors might help in the perception of muscle contraction and body movement has not been settled. The present study gives direct evidence of skin receptor firing in close coincidence with the contraction of the vaginal and anal sphincters. The distal stump of the sectioned motor pudendal nerve was stimulated. Single shocks induced a wavelike increase in the lumen pressure of the distal vagina and the anal canal, as well as constriction of the vaginal introitus and the anus. The constriction pulls on and moves the surrounding skin, which was initially detected visually. In the present experiments, a thin strain gauge that pressed on the skin surface detected its displacement. Single shocks to the motor nerve induced a wave of skin movement with maximal amplitude at 5 mm from the anus and propagated with decrement beyond 35 mm. The peripheral terminals of the sensory pudendal nerve and the posterior femoral nerve supply the skin that moves. Sensory axons from both nerves fired in response to both tactile stimulation and the skin movement produced by the constriction of the orifices (motor-sensory coupling). In cats with all nerves intact, a single shock to the sensory nerves induced reflex waves of skin movement and lumen pressure (sensory-motor coupling). Both couplings provide evidence for a feedforward action that might help to maintain the female posture during mating and to the perception of muscle contraction.

Structure, innervation, mechanical properties and reflex activation of a striated sphincter in the vestibule of the cat vagina.

Vaginal constriction might be important for reproduction in mammals, but existing information is both limited and controversial. This paper shows the structure, mechanical properties, innervation and reflex response of a striated sphincter in the vestibule of the cat vagina. A Foley catheter coupled to a pressure transducer detected in the lumen of the vestibule a pressure wave that was induced by stimulation of the external branch of the motor pudendal nerve. The peak pressure of the wave induced by bilateral stimulation (30.6 cm H(2)O) was about double of the peak pressure wave induced unilaterally. The tetanus/twitch amplitude ratio was 4.5. The sphincter that produces the increase in vaginal pressure fatigues slowly. Digital, point-to-point summation of unilateral waves was greater than the wave induced bilaterally. Summation of the pressure wave induced by the separate stimulation of the terminal motor branches was also greater than the wave induced by the entire motor nerve. This might reflect multiple innervation of muscle fibres. Single, controlled probing of the vaginal vestibule induced a reflex discharge in the motor nerve. Repetitive probing (10 Hz) induced a motor nerve post-discharge lasting >1 min. The vaginal sphincter is two-half rings of striated muscle fibres in the wall of the vaginal vestibule; the fibres end freely in the dorsal and ventral midlines. Penetration of the vestibule by the penis might trigger sustained contraction of the vaginal sphincter.

Pudendal nerve stimulation, interneurons post-discharge and delayed depolarization in hind limb motoneurons of the female cat.

The present experiments were done in the spinal female cat. In a prior work in the decerebrate female cat, stimulation of the sensory pudendal nerve (SPN) induced a depolarizing wave (LD) in hind limb motoneurons that outlasted the stimulus by up to 6 s. LD triggered self-sustained motoneuron firing (bistability). An intrinsic potential underlies bistable firing, which, in the cat, depends on two main factors; first, the integrity of pathways descending from the brain stem to the spinal cord and, second, the membrane potential of the motoneuron just before the stimulus; at high resting potential, excitatory short-lasting inputs induce transient but no sustained firing. Thus, no bistability occurs in the spinal cat or in hyperpolarized motoneurons of the decerebrate cat. LD might be an intrinsic potential that could also be absent in the spinal cat, or an extrinsic (synaptic) potential induced by spinal interneurons. In the latter case, the interneurons generating LD should show post-discharge as prolonged as LD. LD was produced in spinal cats and its amplitude did not change or increase slightly during hyperpolarizing pulses, which suggests that LD might be a synaptic response. Interneurons showing post-discharge to train of stimulation to SPN were located 100-200 microm above the pools of hind limb motoneurons. Some post-discharges were as prolonged as LD. We conclude that LD might be a synaptic response to local interneurons.

Effects of postganglionic nerve section on synaptic transmission in the superior cervical ganglion of the guinea-pig.

In sympathetic neurons, axotomy induces a marked depression in synaptic transmission. We asked whether the decrease in synaptic efficacy observed in a given axotomized cell is more severe if most of the postsynaptic neurons are also injured. Accordingly, we studied if the synaptic depression induced by axotomy in neurons with axons running in a postganglionic nerve is influenced by section of other postganglionic nerves. The excitatory postsynaptic potentials evoked by preganglionic stimulation were recorded intracellularly in an in vitro preparation of the superior cervical ganglion. Eight days after cutting the inferior postganglionic nerve, postsynaptic potentials recorded from neurons projecting through this nerve were smaller (median = 6 mV, n = 62) than the controls (median = 34 mV, n = 89), but were similar to those found after sectioning the inferior nerve plus most postganglionic nerves (median = 5 mV, n = 70). If the inferior nerve was left intact, but most postganglionic branches were cut, the synaptic potentials recorded from inferior nerve neurons were normal (median = 33 mV, n = 77). It is concluded that the synaptic depression induced by axotomy in a sympathetic neuron is not affected by axotomy of nearby ganglion cells, even if they share part of their presynaptic axons. This suggests that the effect of axotomy is restricted to the synaptic terminals on the injured neuron.

Methods to find aponeurosis and tendon stiffness and the onset of muscle contraction.

A method to measure small movements of living tissues either large or small is presented. The method is based on the detection of changes in reflected infrared light. An optocoupler (coupled photodiode and photodetector) and a small (< 1 cm2) mirror were used. The optocoupler (OC) has a low cost and it can be calibrated easily. It can be also used as the transducer of a strain-gage. Three different uses are shown: (a) as a strain-gage transducer; (b) detection of tendon and aponeurosis movements in large muscles (cat soleus); (c) detection of the onset of muscle contraction. Movements of less than 1 microm can be detected with the aid of automatic averaging of the signals. Concerning the second use (b), it permits the estimation of tendon stretch. Concerning the third use, the onset of muscle movement precedes by at least 2 ms that of the force recorded at the tendon.

The influence of peripheral connections on wallerian degeneration.

Following electrophysiological techniques we investigated whether wallerian degeneration of the cat sural nerve may be influenced by (1) its peripheral connection and (2) the distance from the cell body. Distal stumps connected to their innervation sites peripherally showed less degeneration than proximal ones when isolated between two sections. However, when two isolated adjacent segments were produced by making 3 sections, the resulting degeneration was more pronounced in the distal segment. Thus, both the factors mentioned above appear to influence wallerian degeneration.

Inhibition of withdrawal responses by pelvic nerve electrical stimulation.

In urethane-anesthetized rats, the compound action potential of the pelvic nerve was found to consist of three different waves, two in the A delta fiber and one in the C-fiber range of conduction velocity. Electrical stimulation of the pelvic nerve produced a complete inhibition of the withdrawal response to noxious foot pinch or foot compression. The electromyographic (EMG) activity of the contralateral posterior biceps muscle was used to record the withdrawal response. The withdrawal response inhibition was related to the duration and the frequency of electrical stimulation. Low (5-10 Hz) and high (100-300 Hz) frequencies were ineffective in inhibiting the withdrawal response, whereas intermediate frequencies (20-80 Hz) produced a complete inhibition of the withdrawal response. Short (300 ms) trains of stimulation inhibited the withdrawal response only during the stimulation period. Longer trains of stimulation (500 ms-10 s) produced long-lasting inhibition of the response to noxious stimulation. The inhibition persisted for up to 20 s after the end of electrical stimulation of the pelvic nerve. A delta fiber stimulation was adequate to inhibit the withdrawal response in most (15 out of 17) of the animals. However, A delta plus C-fiber stimulation was needed to inhibit the response to noxious stimulation in two animals. In addition to inhibiting the response to noxious stimulation, pelvic nerve electrical stimulation reflexively activated abdominal muscles. On the basis of present findings using electrical stimulation, it can be suggested that, in the rat, A delta and C-visceral afferents of the pelvic nerve mediate the analgesic effect of vaginocervical probing pelvic and A delta afferents the contraction of abdominal muscles in the fetus-expulsion reflex.

The cat pudendal nerve: afferent fibers responding to mechanical stimulation of the perineal skin, the vagina or the uterine cervix.

Some afferent fibers from the pudendal nerve of the female cat were stimulated by pressing on the perineal skin, the vagina or the uterine cervix. Three different types of skin mechanoreceptors were found: (1) with low threshold (< 20 mg) and slow-adapting discharges; (2) with high threshold (0.1-0.5 g) and slow-adapting discharges; and (3) with low threshold and fast-adapting discharges. Most of these receptors increased their firing frequency as the velocity of skin indentation was increased (velocity detectors). The average conduction velocity of the skin afferents was 29 +/- 9 m/s. The receptors located at the vagina showed a fast-adapting response to probing and were sensitive to the velocity of the probe movement. Most of these receptors, however, showed a slow adaptation when the vaginal wall was distended with a balloon. The conduction velocity in vaginal afferents was 37 +/- 16 m/s. Those receptors responding to pressure on the uterine cervix adapted slowly to constant pressure but were sensitive to the velocity of the pressure pulses. The conduction velocity in the afferents from the uterine cervix was 31 +/- 9 m/s.

Prolonged inhibition of the flexor reflex by probing the cervix uteri in the cat.

In decerebrate or spinal cats, sustained mechanical stimulation of the cervix uteri inhibited the flexor reflex elicited by electrical stimulation of the foot pad during the probing period (160 s). After probing, 3-15 min were required for reflex recovery. No additional inhibition was produced if probing was repeated before recovery, but instead the reflex was facilitated. When probing was applied 5-10 min after reflex recovery the reflex was again abolished. The recovery, however, occurred earlier and was followed by facilitation. Probing the cervix with single mechanical pulses inhibited transiently (140-200 ms) the short latency reflex components, but the components with longer latencies are unaffected or facilitated. Distension of the vaginal wall with a balloon also inhibited the flexor reflex, but a transient, mild facilitation appeared several seconds after the distension. In general, whenever the inhibition decreases, the facilitation predominates. Our findings suggest that cervical probing or vaginal distension triggers both a long-lasting inhibition and a concomitant facilitation in different intraspinal flexor reflex pathways.

Sustained activation of the triceps surae muscles produced by mechanical stimulation of the genital tract of the female cat.

In decerebrate cats, controlled mechanical stimulation of the perivulvar skin, the vaginal wall or the cervix uteri induced visible hind limb extension. Pressing on the cervix uteri produced the greater response. To quantify these responses, the EMG activity and the tension developed by the normally inserted triceps surae muscles were recorded. The activity induced in these muscles by stimulation of the genital canal outlasted the stimulus by many seconds or a few minutes. These effects disappeared after spinalization at the T12 level. We propose that stimulation of the vaginal canal in the female cat may induce bistability of triceps surae motoneurones.

Efficacy of a putative GABA analog on synaptic transmission in the cat spinal cord.

4-Hydroxy-4-phenylcaproamide (HPhCA), at high doses or rates of IV injection depressed the ventral root reflexes elicited by nerve or dorsal root stimulation. The D (direct) and I (synaptic) ventral root waves and the antidromic (A) dorsal root wave evoked by intraspinal stimulation were also depressed. Similar effects were produced when HPhCA was applied topically on the cord dorsum. At 80 mg/kg and 8 mg/kg/min, the spinal reflexes and the I wave were facilitated for 4 to 6 h, but the D and A waves were depressed. Intracellular recordings from motoneurons showed that HPhCA injection produced: hyperpolarization that lasted several hours, short lasting (< 20 min) facilitation of both EPSPs and IPSPs as well as spike-like potentials (SLPs) that were triggered by EPSPs even though the neuron was hyperpolarized. SLPs may reach the threshold for full spikes. Our results suggest that the spinal depression results from hyperpolarization of motoneurons and the initial facilitation appears to be presynaptic. The late facilitation may be produced by SLPs. HPhCA does not appear to mimic the actions of GABA in primary afferents fibers and motoneurons.

Spectrophotometric and chromatographic detection of Karwinskia humboldtiana (tullidora) toxin in rat serum after tullidora ingestion.

The absorbance spectrum of tullidinol, a tullidora (Karwinskia humboldtiana) toxin, showed peaks at 260 and 410 nm when dissolved in ethanol; the second peak was linearly related to toxin concentration. Silica gel thin layer chromatograms of tullidinol solutions showed a single, yellow spot with an Rf of 0.76. A single dose of tullidora fruit homogenate was orally given to male Wistar rats; the absorbance spectrum of serum from tullidora-treated rats showed peaks at 300 nm and at 410 nm; the second peak was absent in serum from control rats. No spots were detected in chromatograms of normal serum, but a yellow spot with the same Rf as that of tullidinol was found in the serum of tullidora-treated rats. We conclude that tullidinol was detected in the serum of treated animals and that the methods we used for its detection may also help to diagnose accidental tullidora poisoning in cattle and human beings.

Effects of activity in single sensory fibres on the discharge patterns of dorsal spinocerebellar tract cells.

1. Electrical activity of dorsal spinocerebellar tract (DSCT) was recorded extracellularly and intracellularly in Clarke's column nucleus. 2.Trains of impluses were elicited in DSCT cells by static stretch of the hind limb muscles, and the distributions of intervals between the impulses were computed. These distributions were essentially similar before and after impalement of the cells with the micropipette, suggesting that cell injury caused by penetration was insignificant. 3. Patterns of synaptic activity revealed that the DSCT cells examined were innervated by a limited number of muscle afferents. 4. In some DSCT neurons, all excitatory post-synaptic potentials (e.p.s.p.s) elicited by a single fibre were suprathreshold, and e.p.s.p.s elicited by other afferents were usually subthreshold. These neurons discharge very regularly during muscle stretch, and the spike initiation followed nearly one-to-one relation in response to activity of the sensory fibre eliciting suprathreshold e.p.s.p.s. 5. The discharge pattern was irregular in other DSCT cells. In these cases, single sensory fibres evoked e.p.s.p.s. which did not always reach the firing threshold because of random fluctuation in their amplitude. 6. DSCT neurons often show low frequency discharges (about 10/sec) in the absence of sensory activation. In these conditions only small e.p.s.p.s. were detected (average, 0.53 mV) suggesting that the average membrane potential is very close (1-i mV) to the firing threshold. 7. It is concluded that the statistical properties of firing patterns in DSCT neurons activated by muscle afferents depend on (a) the large amplitude of e.p.s.p.s., (b) their firing threshold and (c) random fluctuations in the e.p.s.p.s. amplitude.

Quantitative changes in myelin proteins in a peripheral neuropathy caused by Tullidora (Karwinskia humboldtiana).

Peripheral nerve demyelination was induced in cats by oral administration of ether extracts of Tullidora (Karwinskia humboldtiana). Proteins from several hindlimb nerves, spinal roots, and dorsal columns of the spinal cord were subjected to slab gel electrophoresis and quantified by densitometry. In Tullidora-treated cats with severe motor disturbances, specific myelin proteins were reduced by at least 50% in motor nerves and less than 25% in cutaneous axons. There was a greater decrease of these proteins in the distal than in the cephalad segments of the sciatic nerve; no changes were detected either in the spinal roots or in the white matter of the spinal cord. Electron microscopy revealed intense demyelination in the motor nerves only. Both the density of the 100 A-thick neurofilaments and the relative proportion of a polypeptide with a molecular weight of 68,000 were considerably increased in the affected nerves. It is tentatively concluded that the active principles of Tullidora may enter the axons through the motor nerve terminals. The distal segments of the motor nerves would then be preferentially affected and demyelination could result from axonal damage.

Distal reduction of the conduction velocity of alpha-axons in tullidora (buckthorn) neuropathy.

Cats were given a single oral dose of ether extracts of the tullidora (Karwinskia humboldtiana) fruit. Three to five weeks later these animals developed the tullidora (buckthorn) neuropathy. Acute experiments were conducted in control cats as well as in treated cats which showed hind limb paralysis. Spinal motoneurons of the medial gastrocnemious and the soleus nerves were recorded intracellularly and their axons were electrically stimulated at two points, near the entry to the muscle and at the hip level. The conduction velocity was determined for the segment between the two stimulating electrodes [distal conduction velocity; (DCV)] and for the segment between the electrode at the hip level and the spinal cord [proximal conduction velocity (PCV)]. In untreated animals, DCV was higher than PCV so that the ratio DCV:PCV was, on the average, larger than 1. In treated animals, DCV was reduced, but PCV remained within normal limits and DCV:PCV was lower than 1. In addition, there was a relative deficit of the faster velocities suggesting that the thicker fibers were preferentially affected. These observations are in agreement with previous ones indicating that demyelination induced by tullidora is restricted to distal nerve segments.

Tullidora (Karwinskia humboldtiana) toxin mainly affects fast conducting axons.

Cats were given a single oral dose of ether extracts from tullidora (Karwinskia humboldtiana) fruit which contains an identified neurotoxin. Acute experiments were performed 4-7 weeks after toxin administration when flaccid limb paralysis was evident. Normal cats were used as controls. The medial gastrocnemius, the soleus and the sural nerves were electrically stimulated and the unitary potentials evoked by the stimuli were extracellularly recorded from spinal root filaments to measure the conduction velocity of single fibres. In control cats, the average conduction velocity (CV) was greater in medial gastrocnemius motor fibres than in the afferent ones of the same nerve and the soleus motor axons, whereas in the sural nerve CV was less than in the aforementioned cases. The CV values and the proportion of fast conducting fibres (greater than 80 m/s) in each nerve were directly related (r = 0.99). In treated cats, CV diminished in all the nerves studied, but the conduction velocity was further reduced in the faster fibres. Consequently, the motor division of the medial gastrocnemius nerve, normally composed of a high proportion (57%) of fast fibres, was more affected by tullidora and the sural nerve, which has the lowest proportion (0.7%) of these type of fibres, was the less affected. Our findings suggest that the preferential involvement of motor nerves in the experimental tullidora (buckthorn) neuropathy, as well as the preservation of somatic sensation in quadriplegic children accidentally poisoned with tullidora, are related to the distribution of axonal diameters in peripheral nerves.

Axonal transport: a quantitative study of retained and transported protein fraction in the cat.

The fast axonal transport of proteins was studied in the cat sciatic nerve after injection of [3H]leucine into the spinal ganglion or the ventral horn of the seventh lumbar segment. The amount of transported proteins after ganglion injection was linearly related to the amount of label present at the ganglion. At variable intervals after ganglion or spinal cord injection, the sciatic nerves were sectioned in some experiments. The transport of proteins continued in the peripheral nerve stump in a wavelike manner, but the advancing wave leaves a labeled trail behind. A fraction of this trail corresponds to proteins moving at slower velocities than the velocity of proteins in the wave front. Another fraction of the trail corresponds to molecules retained by the axons. Each nerve segment of 5 mm in length retains 1.5% of the transported proteins, and the profile of retained proteins along the sciatic nerves follows a single exponential function. From the proportion of retained proteins, the concentration of transported proteins at the terminals of branching axons as a function of the branching ratio was estimated. In the case of motor axons innervating the soleus muscle of the cat, the concentration of recently transported proteins at the nerve terminals would be approximately 0.83% of the proteins leaving the spinal cord. This low concentration of transported proteins at the nerve terminals may explain the lability of neuromuscular synapses when axonal transport is decreased or interrupted.

Serial dependency in the discharge pattern of dorsal spinocerebellar tract neurons: a computer simulation analysis.

We have developed a model in order to analyze the factors eventually responsible for the strong negative serial dependency between successive interspike intervals in the discharge of the Dorsal Spinocerebellar Tract (DSCT) neurons. This dependency is reflected, phenomenologically, by short intervals followed by long ones and, quantitatively, by the first order correlation coefficient (R1-2); which can be lower than -.6 (Jansen, Nicolaysen & Rudjord, 1966; Kröller and Grüsser, 1982). We have found that the lowest values of R1-2 are always related with model parameter values which were very similar to those obtained experimentally. It was observed that EPSP amplitude distribution plays an important role in the discharge patterns of the DSCT neurons. There is one fiber that elicits EPSPs greater than 6 mV, which is responsible for the genesis of the short intervals in the discharge. Long intervals are determined basically by a suprathreshold depolarization and the afterhyperpolarization processes.

Axon-to-axon transmission in tullidora (buckthorn) neuropathy.

Oral administration of ether extracts of the tullidora ( Karwinskia humboldtiana ) fruit, which contains an identified neurotoxin, produced flaccid hind limb paralysis in cats after a latency of 4 to 7 weeks. Acute experiments were conducted after the paralysis was evident. Spinal roots of lumbar and sacral segments were transected as close as possible to the spinal cord and divided into several filaments. Stimulation of some filaments distal to the transection evoked action potentials in other filaments (axon-to-axon transmission or cross talk) after a latency of at least 8 ms. Cross-talk responses frequently consisted of multiple discharges. Axon-to-axon transmission was seen only between motor axons and disappeared when hind limb nerves were transected 10 to 15 cm from the spinal cord. Twin pulses were applied to a filament at various intervals; the pulse intensity was adjusted so that the conditioning pulse was subthreshold to elicit cross talk, but the test pulse frequently elicited it (temporal facilitation). In three fully studied fibers the facilitation was prolonged to 50 to 80 ms. In some cases, no cross talk was evoked in a given filament by individual stimulation of two other filaments, but simultaneous stimulation of the same filaments did evoke cross talk (spatial facilitation). Series of periodic bursts of activity spontaneously occurred in those axons responding with multiple discharges to single stimulation of other axons. At low temperatures (about 30 degrees C) the stimulus could trigger essentially similar series of bursts. Single motoneurons were intracellularly stimulated by brief depolarizing pulses. The action potential elicited by the stimulus was followed after several msec by a secondary train of discharges generated at the periphery ("back firing").

Denervation caused by tullidora (Karwinskia humboldtiana).

Crude homogenate of the seed of the tullidora (Coyotillo, buckthorn; Karwinskia humboldtiana) fruit or a purified neurotoxin extracted from it was administered orally to male Wistar rats. Three to 5 weeks later, gait disturbances and progressive flaccid paralysis of the hindlimbs appeared in these animals which were then submitted to acute experiments. Single twitch and tetanic contractions of the soleus muscle were studied in the tullidora treated rats. The twitch tension elicited by direct stimulation of the muscle was greater than that produced by nerve stimulation. In addition, the tension of nerve-elicited twitches diminished as the stimulating electrode was moved towards the spinal cord, suggesting failure of nerve conduction. When the muscle was directly stimulated, contraction and relaxation were slower than normal and the tetanus/twitch tension ratio was subnormal. These data suggest partial denervation of the soleus muscle under the action of the toxin. Electron microscopy revealed abnormal motor-nerve endings with few synaptic vesicles and denervated end-plates. Degenerative changes of axons and myelin disturbances were frequently seen in the intramuscular branches of the soleus nerve. However, alterations of this type were less severe in the nerve sections closer to the spinal cord and no significant changes were detected at the upper portion of the sciatic nerve. It is concluded that the paralysis produced by the tullidora toxin is the result of both conduction block of the nerve impulses and muscle denervation.