Influences of external urethral sphincter relaxation induced by alpha-bungarotoxin, a neuromuscular junction blocking agent, on voiding dysfunction in the rat with spinal cord injury.

OBJECTIVES: To determine whether external urethral sphincter (EUS) relaxation induced by alpha-bungarotoxin, a highly selective neuromuscular junction blocking agent, could ameliorate voiding dysfunction after spinal cord injury (SCI) in rats. METHODS: The effects of intravenous alpha-bungarotoxin (333 microg/kg) were evaluated during cystometry in decerebrate, unanesthetized female Sprague-Dawley rats (250 to 300 g) with spinal cords chronically transected at T7-9 (n = 7) or with normal spinal cords (NSC) (n = 7). Parameters measured included voided volume (VV), residual volume (RV), volume threshold for inducing micturition (VT), voiding efficiency (VE), micturition pressure (MP), pressure threshold for inducing micturition (PT), bladder contraction duration (BCD), and compliance (CP). RESULTS: In SCI rats, treatment with alpha-bungarotoxin improved voiding. The toxin increased VE (31%) and reduced RV (42%), MP (52%), BCD (14%), and VT (31%). VV, PT, and CP were not altered. In NSC rats, alpha-bungarotoxin decreased VE (23%), increased RV (63%), and decreased MP (36%), VV (38%), and VT (20%) but did not change BCD and CP. CONCLUSIONS: The results of our study demonstrated that alpha-bungarotoxin improved voiding in SCI rats by reducing urethral outlet resistance. However, in NSC rats, the toxin reduced voiding, probably by suppressing high-frequency phasic sphincter activity, necessary for efficient urine elimination in normal animals. The present results provide further support for the view that drugs that depress striated muscle activity can be useful in the treatment of voiding dysfunction after SCI.

Plasticity in spinal opioid control of lower urinary tract function in paraplegic cats.

Spinal cord injury disrupts micturition reflexes, which produces morbidity. The contribution of endogenous opioid systems to urinary retention were assessed in chronic spinal cats by administering the opioid receptor antagonist, naloxone (5-500 micrograms kg-1, i.p.), to unanesthetized paraplegic cats while monitoring lower urinary tract function and observing hind limb reflexes. While naloxone had no overt effect in acute spinal cats, in chronic spinal cats naloxone induced the release of large volumes of urine and produced marked hind limb hyper-reflexia. Prominent tachyphylaxis and tolerance to the effects of naloxone were evident. Immunohistochemical studies indicated a marked increase in leucine enkephalin and dynorphin in sacral spinal neurons. Together, these data indicate hyperactivity of the endogenous spinal opioid system following recovery from spinal cord injury and, furthermore, suggest that the spinal neural circuitry may become 'dependent' upon elevated levels of endogenous opioid peptides.

Selective inhibitory effects of ethylketocyclazocine on reflex pathways to the external urethral sphincter of the cat.

In the ventral horn of the sacral spinal cord of the cat, opioid terminals are preferentially localized in Onuf's nucleus, an area containing motor neurons that innervate the striated muscle of the external urethral sphincter. The present study was undertaken to 1) compare the effects of selective opioid agonists on sphincter reflex pathways with the effects of these drugs on hindlimb reflexes and urinary bladder reflexes and 2) determine if the physiological inhibition of sphincter reflexes, which accompany bladder contractions, is mediated by endogenous opioids. The effects of intrathecal (i.t.) and i.v. drug administration on bladder activity, sphincter reflexes and reflexes to the hindlimb musculature were monitored in chloralose-anesthetized cats. Ethylketocyclazocine (0.05-500 micrograms i.t.) produced a dose-dependent, naloxone-sensitive, inhibition of sphincter reflexes to less than 10% of control amplitude while having no consistent effects on hindlimb reflexes or bladder activity. D-Ser2-leu5-enkephalin-thr6 (DSLET; 0.1-2.0 micrograms i.t.) abolished rhythmic bladder activity, while having no effects on sphincter or hindlimb reflexes. Larger doses of DSLET (5.0-10 micrograms i.t.) produced a modest reduction of sphincter reflexes (to 60% of control amplitude), without affecting hindlimb reflexes. Naloxone (50 micrograms i.t.) reversed DSLETs marked inhibition of bladder activity, whereas large doses (greater than 250 micrograms i.t.) only partially antagonized DSLETs weak inhibition of sphincter reflexes. Morphine (5-500 micrograms i.t.) had no consistent effect on any of the measures.(ABSTRACT TRUNCATED AT 250 WORDS)

Sympathetic innervation of the cornea from the superior cervical ganglion. An HRP study in the cat.

Horseradish peroxidase (HRP) was applied to the cornea in cats using a variety of methods. Small numbers of labeled neurons were observed in the superior cervical ganglion (SCG), the majority were located in the rostral half. This confirms previous histofluorescence data which indicated the SCG was the likely source of adrenergic fibers to the cornea.

Organization of corneal afferent axons in the trigeminal nerve root entry zone in the cat.

Experiments were carried out in cats to learn the location of sensory axons from the cornea in the trigeminal nerve root just prior to its entry into the brainstem. HRP injected into the cornea labelled these axons and indicated they were not restricted to the ophthalmic division of the nerve root as had been indicated from previous studies. These findings, if representative of other branches of this nerve, offer a partial explanation for the variable preservation of function following transection of an entire division of the trigeminal nerve root in cases of trigeminal neuralgia.

The distribution within the spinal cord of visceral primary afferent axons carried by the lumbar colonic nerve of the cat.

Horseradish peroxidase taken up by the sensory axons in the lumbar colonic nerves in 5 cats was observed in the dorsal root ganglia and in the spinal cord in segments L1 through L5. Reaction product was observed in Lissauer's tract, the dorsal columns and laminae I, V, VII and X in a pattern typical of visceral primary afferents from other nerves. A small number of preganglionic neurons were also labeled.

The spinal distribution of sympathetic preganglionic and visceral primary afferent neurons that send axons into the hypogastric nerves of the cat.

The spinal distribution of sympathetic preganglionic neurons (PGN) and visceral primary afferent neurons sending axons into the hypogastric nerve of the cat has been studied with HRP tracing techniques. After application of HRP to the cat hypogastric nerve, labeled PGN were identified in segments L2-L5. Most of these neurons were oriented transversely and were divided approximately equally between two nuclei: the principal nucleus and the intercalated nucleus. Cells were distributed in clusters at 160-361-microns intervals along the length of the cord. Sensory neurons were labeled in dorsal root ganglia from T12 to L5. Central axons of these visceral afferents were observed in the medial half of Lissauer's tract from T13 to L7. Afferent axon collaterals extended through lamina I on both sides of the dorsal horn but were most prominent on the lateral side, where they continued into lateral lamina V and VII, often overlapping the dorsal dendrites of PGN in this region. Labeled afferent projections exhibited a periodic distribution in lamina I with clusters of axons occurring at 235-343-microns intervals in the rostrocaudal axis. The central projection of hypogastric nerve primary afferents was qualitatively similar to the distribution of visceral afferent projections at other levels of the spinal cord.

Tracing of afferent and efferent pathways in the left inferior cardiac nerve of the cat using retrograde and transganglionic transport of horseradish peroxidase.

Retrograde and transganglionic transport of horseradish peroxidase (HRP) was used to trace afferent and efferent pathways in the left inferior cardiac nerve of the cat. Cardiac efferent and afferent neurons were located, respectively, in the stellate ganglion (average cell count per experiment:2679) and in the ipsilateral dorsal root ganglia (DRG) from C8 to T9 (average cell count per experiment:213). Labeled cardiac afferent projections to the spinal cord were most dense in segments T2-T6 where they were located in Lissauer's tract and in lamina 1 on the lateral border of the dorsal horn. Labeled afferent axons extended ventrally through lamina 1 into lamina 5 and the dorsolateral region of lamina 7 in proximity to the intermediolateral nucleus. A weak projection was noted on the medial side of the dorsal horn. These sites of termination are similar to projections by other sympathetic afferent pathways (i.e. renal, hypogastric and splanchnic nerves) to the lower thoracic and lumbar spinal cord, indicating that visceral afferents may have a uniform pattern of termination at various segmental levels. This pattern of termination in regions of the gray matter containing spinothalamic tract neurons and neurons involved in autonomic mechanisms is consistent with the known functions of sympathetic afferent pathways in nociception and in the initiation of autonomic reflexes.

Selective inhibition of peripheral cholinergic transmission in the cat produced by AF64A.

The neuropharmacologic effects of ethylcholine aziridinium ion, AF64A, were studied in cats, using various physiological techniques, to ascertain its synaptic site of action and to determine whether it may act as a cholinergic specific neurotoxin in vivo. Nictitating membrane contractions elicited by preganglionic nerve stimulation (1-16 Hz) were diminished in a dose-dependent manner after injection of AF64A into the carotid artery. Contractions due to injection of l-norepinephrine, tetramethylammonium or acetylcholine were not changed. Postganglionic action potentials from the superior cervical ganglion evoked by preganglionic stimulation were also abolished by AF64A, whereas the postganglionic firing produced by tetramethylammonium was unchanged. Neither the nictitating membrane nor ganglionic responses on the contralateral side of the animal were affected by AF64A treatment. In the same animals, twitch tension in the tongue produced by stimulation of the ipsilateral hypoglossal nerve (1-16 Hz) was gradually reduced and in most experiments completely blocked by AF64A. Repetitive stimulation of either the autonomic or somatic nerves at high frequencies (greater than 10 Hz) magnified and accelerated the onset of neurotoxic effects of AF64A. The suppression of ganglionic and neuromuscular transmission by AF64A was irreversible during the course of the experiments (12-18 hr). From these results, we can conclude that AF64A produces in the peripheral nervous system a longlasting inhibition of cholinergic transmission, without interfering with adrenergic transmission. Moreover, because AF64A did not block the postganglionic responses elicited by cholinergic nicotinic and muscarinic agonists, the inhibitory effects of AF64A must be mediated by a presynaptic action on cholinergic nerve terminals.

Naloxone induced micturition in unanesthetized paraplegic cats.

In chronic spinal cats 2 to 10 weeks after transection of the spinal cord at the lower thoracic level (T12-T13), the administration of naloxone, an opiate antagonist, (32-500 micrograms./kg. i.p.), stimulated micturition. The total quantity of urine released after administration of naloxone ranged from 10 to 70 per cent, (mean 39 per cent) of the initial bladder volume. The response to the drug occurred 5 to 10 minutes after injection and was characterized by repeated periodic expulsion of small quantities of urine (5 to 10 ml.) which coincided with a pattern of hind-limb movement which resembled walking behavior. The effects of naloxone persisted for about 1 hour. The motor activity following administration of naloxone was dependent upon activation of bladder afferents since it did not occur when the bladder was empty. Naloxone also facilitated the release of urine induced by stimulation of somatic afferents. With repeated administration of naloxone, tolerance developed which was evident for several days. These observations suggest that an endogenous opiate may have a tonic inhibitory role in regulation of micturition. Pharmacologic manipulation of this putative inhibitory mechanism may facilitate management of neurogenic bladder dysfunction.

Location and morphology of parasympathetic preganglionic neurons in the sacral spinal cord of the cat revealed by retrograde axonal transport of horseradish peroxidase.

The distribution and morphology of preganglionic neurons in the sacral parasympathetic nucleus (SPN) of the cat have been studied with the horseradish peroxidase (HRP) tracing technique. HRP applied to the cut pelvic nerve was identified in cells located ipsilaterally, primarily in the intermediate gray matter. They formed a column approximately 10 mm long, usually contained within two, but occasionally three, sacral segments, S2 contained a majority of cells. In transverse sections the SPN had the appearance of an inverted "L". Cells were medium-sized, oval or spindle-shaped, and transversely oriented. They were distributed among two major components and one minor one: (1) dorsal band (34%) located mainly in lamina V beneath the dorsal horn (cells and dendrites horizontally oriented), (2) lateral band (64%) along the lateral edge of the gray matter in laminae VII through V (cells oriented dorsoventrally with dendrites extending within the nucleus and into the dorsolateral funiculus), and (3) a small group (2%) of cells at the rostral end of the SPN in lamina VII in the middle of the ventral horn. These data coupled with the results of other investigations indicate that the SPN has a viscerotopic organization wherein the colon is innervated primarily by cells in the dorsal band and the urinary bladder is innervated primarily by cells in the lateral band.

Physiology of the urinary bladder and urethra.

Activation of the parasympathetic pathways to the detrusor muscle and inhibition of somatic input to the external urethral sphincter are the essential neuronal events initiating release of urine. The former occurs via a spinobulbospinal pathway, whereas the latter is produced by inhibitory mechanisms in the sacral spinal cord. The sympathetic outflow to the urinary tract promotes urine storage by increasing urethral resistance and depressing detrusor contractions. Sympathetic activity is generated at least in part by a spinal vesicosympathetic reflex pathway. Evidence indicates that the integration of sympathetic and parasympathetic inputs to the bladder can occur at the level of the peripheral autonomic ganglion as well as at levels of the effector organ. The existence of facilitatory and adrenergic inhibitory mechanisms in ganglia and the identification of spontaneously active ganglion cells raise the possibility that vesical ganglia may have a role in modulating or "filtering" the efferent neural input to the bladder.

Physiology of male sexual function.

The male sexual response cycle consists of excitement, plateau, orgasm, and resolution. The initial event, penile erection, is produced by arteriolar dilatation and increased blood flow to the erectile tissue of the penis. Erection is a reflex response initiated by visual, olfactory, or imaginative stimuli impinging upon supraspinal centers or by genital stimulation that in turn activates spinal reflex mechanisms. Sacral parasympathetic and thoracolumbar sympathetic nerves provide the efferent vasodilator input to the penis. Parasympathetic nerves also stimulate secretion from the seminal vesicles and prostate and Cowper's glands during the plateau phase. The orgasmic phase is characterized by seminal emission and ejaculation and the accompanying sensations. Emission of semen into the urethra depends on sympathetic nerves that elicit contractions of smooth muscles in the vas deferens, seminal vesicles, and prostate. Rhythmic contractions of striated muscle (bulbocavernosus and ischiocavernosus) generated by efferent pathways in the pudendal nerve eject semen from the urethra.

The central origin of efferent pathways in the carotid sinus nerve of the cat.

The application of horseradish peroxidase to the central cut end of the carotid sinus nerve of the cat produced retrograde labeling of neurons in the ipsilateral medulla in the region of the nucleus ambiguus at anterior-posterior coordinates -8 to -10.5. These data coupled with previous electrophysiological observations suggest that the nucleus ambiguus may be the origin of an efferent inhibitory pathway to the carotid body.

Synaptic transmission in parasympathetic ganglia in the urinary bladder of the cat.

1. Electrophysiological techniques were used to study the sacral para-sympathetic input to pelvic ganglia located on the surface of the urinary bladder of the cat. 2. Synaptic transmission in pelvic ganglia was mediated primarily via nicotinic receptors although muscarinic excitatory receptors were present. 3. The most prominent characteristic of transmission in pelvic ganglia was the marked recruitment elicited by increasing frequencies of preganglionic nerve stimulation. Post-ganglionic action potentials were of low amplitude at low frequencies of stimulation (0-1-0-5c/s), but commonly increased to five to twenty times control amplitudes during continuous stimulation at frequencies between 5 and 10c/s. Thus, it is proposed that vesical ganglia may act as "filters" in the micturition pathway; blocking the excitatory input to the bladder when intravesical pressure and parasympathetic firing is low and facilitating the neural input to the bladder during micturition when preganglionic activity is high. 4. Information was also obtained about the characteristics of the parasympathetic post-ganglionic neurones innervating the bladder. Stimulation of the preganglionic fibres in the pelvic nerve elicited a bimodal contraction consisting of an initial phasic response, which was atropine-resistant and a tonic response which was blocked by atropine. This suggests that two types of neurones, cholinergic and non-cholinergic, may mediate the sacral input to the vesical smooth muscle.