A.M. Booth

Organization of the sacral parasympathetic reflex pathways to the urinary bladder and large intestine.

Electrophysiological and horseradish peroxidase (HRP) techniques have provided new insights into the organization of the sacral parasympathetic reflex pathways to the large intestine and urinary bladder. The innervation of the two organs arises from separate groups of sacral preganglionic cells: (1) a dorsal band of cells in laminae V and VI providing an input to the intestine; and (2) a lateral band of cells in lamina VII providing an input to the bladder. These two groups of cells were separated by an interband region containing tract cells and interneurons. Neurons in the interband region received a visceral afferent input and exhibited firing correlated with the activity of intestine and urinary bladder. It seems reasonable therefore to consider the interband region as a third component of the sacral parasympathetic nucleus. Anterograde transport of HRP revealed that visceral afferents from the intestine and bladder projected into the parasympathetic nucleus. Most of the projections were collaterals from afferent axons in Lissauers tract that passed in lamina I laterally and medially around the dorsal horn. These afferent collaterals were located in close proximity to preganglionic perikarya and dendrites in laminae I, V and VI. The proximity of visceral afferents and efferents in the sacral cord probably reflects the existence of polysynaptic rather than monosynaptic connections since electrophysiological studies revealed that both the defecation and micturition reflexes occurred with very long central delays (45-70 msec). The reflex pathways mediating defecation and micturition in cats with an intact neuraxis were markedly different. Defecation was dependent upon a spinal reflex with unmyelinated (C-fiber) peripheral afferent and efferent limbs. On the other hand, micturition was mediated by a spinobulbospinal pathway with myelinated peripheral afferent (A-fiber) and efferent axons (B-fiber). Transection of the spinal cord at T12-L2 blocked the micturition reflex but only transiently depressed the defecation reflex. In chronic spinal cats the micturition reflex recovered 1-2 weeks after spinalization; however, in these animals bladder-to-bladder micturition reflexes were elicited by C-fiber rather than A-fiber afferents. The C-fiber afferent-evoked reflex was weak or undetectable in animals with an intact neuraxis. Transection of the spinal cord also changed the micturition reflex in neonatal kittens (age 5-28 days). In neonates with an intact neuraxis bladder-to-bladder reflexes occurred via a long latency spinobulbospinal pathway (325-430 msec). The long latency is attributable to the slow conduction velocity in immature unmyelinated peripheral and central axons. In chronic spinal kittens (3-7 days after spinalization) the long latency reflex was abolished and a shorter latency (90-150 msec) bladder reflex was unmasked. The emergence of this spinal pathway may reflect axonal sprouting and the formation of new reflex connections within the sacral parasympathetic nucleus.

Parasympathetic preganglionic neurons in the sacral spinal cord

Two types of preganglionic neurons have been identified in the sacral parasympathetic nucleus (SPN) of the cat. These neurons could be differentiated by various characteristics including axonal conduction velocities, morphology, location in the nucleus, organ of innervation and central reflex mechanisms controlling their activity. Neurons having myelinated axons (B-PGNs) with conduction velocities between 3.3 and 13 m/s were located in the lateral band of the SPN and innervated the urinary bladder. Neurons with unmyelinated axons (C-PGNs) with conduction velocities of 0.5-1.4 m/s were located in the dorsal band of the nucleus and innervated the large intestine. B-PGNs were excited by distention of the bladder and inhibited by distension or mechanical stimulation of the intestine, whereas C-PGNs exhibited the opposite responses to these stimuli. C-PGNs often exhibited a low level of spontaneous discharge in absence of stimulation but exhibited marked firing (3.5-10 spikes/s) during a defecation reflex elicited by mechanical stimulation of the rectum-anal canal. The excitatory responses were elicited by C-fiber afferents via a spinal reflex pathway. B-PGNs were inactive when intravesical pressure was below the threshold for inducing micturition (5 cm H2O) but raising the pressure above the threshold induced firing consisting of repetitive bursts of action potentials occurring at relatively high frequencies (15-60 spikes/s). These bursts coincided with rhythmic bladder contractions. The frequency of bladder contractions and associated bursts of PGN-firing and the mean PGN-firing rate (2-8 spikes/s) increased as intravesical pressure was increased in steps between 5 and 30 cm H2O. However, as indicated by interspike interval histograms, the frequency of firing within a burst of action potentials was unchanged. It is concluded that the micturition reflex pathway is organized as a simple on-off switching circuit and that B-PGNs receive a maximal synaptic input when intravesical pressure exceeds the micturition threshold. This circuit was triggered by vesical A delta afferents via a spinobulbospinal pathway. Transection of the spinal cord interrupted the reflex pathway and blocked micturition. However, in chronic spinal animals a spinal reflex mechanisms emerged which contributed to the recovery of bladder function. This mechanism, which was weak or non-existent in animals with an intact neuraxis, exhibited a number of important differences from the normal micturition reflex, most notably being activated by a C-fiber afferent rather than a A delta afferent limb. The mechanism underlying the emergence of C-fiber evoked bladder reflexes in spinal animals is uncertain.

The role of neuropeptides in the sacral autonomic reflex pathways of the cat.

Immunohistochemical and pharmacological studies were conducted to examine the origin and function of peptidergic nerves in the sacral autonomic system of the cat. Leucine-enkephalin (L-Enk) immunoreactivity was identified in nerve terminals in peripheral ganglia on the surface of the urinary bladder and in the parasympathetic nucleus in the sacral spinal cord. In colchicine-treated animals L-Enk was also detected in sacral preganglionic neurons (sPGN) identified by retrograde transport of a fluorescent dye. L-Enk terminals in bladder ganglia are believed to arise from sPGN since the terminals were eliminated by transection of the sacral ventral roots. Pharmacological studies indicated that exogenous as well as endogenously released enkephalins have an inhibitory action at both ganglionic and spinal sites in the sacral outflow to the urinary bladder. Peptides were also associated with afferents nerves in the sacral autonomic system. The distribution of substance P, VIP and cholecystokinin in the sacral dorsal horn paralleled the distribution of visceral afferent projections as demonstrated with HRP techniques. Dye labeling combined with immunohistochemistry revealed that some dorsal root ganglion cells projecting to the pelvic viscera contain substance P or VIP.

Distribution of neurons in the major pelvic ganglion of the rat which supply the bladder, colon or penis.

SummaryIn male rats a large number of the postganglionic neurons which innervate the pelvic organs are located in the major pelvic ganglion. In the present study we have identified the location within this ganglion of neurons which project to either of three pelvic organs, the penis, colon or urinary bladder. Two fluorescent retrogradely-transported dyes, Fast Blue and Fluoro-Gold, were used. For most animals one dye was injected into the cavernous space of the penis, the wall of the distal colon or the wall of the urinary bladder. In a small number of animals two organs were injected, each with a different dye. One to six weeks after injection the major pelvic ganglia were fixed in buffered formaldehyde. The distribution of fluorescent dye-labelled cells was observed in whole mounts of complete ganglia and, in most cases, also in small accessory ganglia located between the ureter and the prostate. The studies showed a unique pattern of distribution for each organ-specific group of neurons. Most of the colon neurons are located in the major pelvic ganglion near the entrance of the pelvic nerve, whereas almost all of the penis neurons are near or within the penile nerve. Bladder neurons are relatively evenly distributed throughout the ganglion. These results demonstrate a distinct topographical organization of organ-specific neurons of the major pelvic ganglion of the male rat, a phenomenon which has also been observed in other peripheral ganglia.

Morphological and electrophysiological properties of pelvic ganglion cells in the rat

Intracellular recording and dye injection were used to study the morphological and electrophysiological properties of rat pelvic ganglion cells. The dye-injected cells measured on the average 37 micron by 22.5 micron and had a mean number of 1.5 primary processes (axon and dendrites). The cells received unmyelinated preganglionic inputs from either the pelvic (parasympathetic) or the hypogastric (sympathetic) nerves, but no cells received inputs from both nerves. The number of synaptic inputs to each cell varied between 1 and 5 with a mean of 2. Each cell had at least one large amplitude suprathreshold EPSP which always initiated an action potential. These properties, namely, morphological simplicity, small number of inputs, security of synaptic transmission and lack of convergence between sympathetic and parasympathetic inputs, suggest that the capacity for synaptic modulation and integration in this ganglion is minimal. Such a structure should therefore relay preganglionic information to target organs with little or no alteration.

The effects of naloxone on the neural control of the urinary bladder of the cat

Naloxone in doses ranging from 0.5 to 512 micrograms/kg i.v., enhanced reflex contractions of the urinary bladder of the cat. At the lowest doses (threshold, 0.5-5 micrograms/kg) the drug increased the frequency of spontaneous bladder contractions. In large doses (10-100 micrograms/kg) the drug produced an initial tonic contraction of bladder lasting 15-40 min followed by a period of high frequency rhythmic activity. Multiunit firing in parasympathetic postganglionic nerves on the surface of the urinary bladder was also enhanced. Bursts of firing which in untreated animals occurred during large bladder contractions occurred continuously during the entire sustained contraction of the bladder following large doses of naloxone. Various evidence indicates that the site of action of naloxone is in the central nervous system. These findings suggest that the parasympathetic reflex pathway to the urinary bladder may be subject to tonic enkephalinergic inhibitory control.

The presence of leucine-enkephalin in the sacral preganglionic pathway to the urinary bladder of the cat ☆

Leucine-enkephalin (L-ENK) nerve terminals which surround the cholinergic neurons in ganglia of the cat urinary bladder are eliminated after transection of the sacral ventral roots or the pelvic nerve. These findings, coupled with other anatomical and physiological data, suggest that L-ENK may be a cotransmitter with acetylcholine in the sacral preganglionic pathways to the urinary bladder.

Regulation of Urinary Bladder Capacity by Endogenous Opioid Peptides

Naloxone administered to chloralose or ketamine anesthetized cats reduced urinary bladder capacity. Successive cystometrograms revealed that naloxone in doses of 0.5 microgram./kg. to 15 micrograms./kg. i.v. reduced the volume necessary to evoke micturition by 10 to 50 per cent, respectively. The effect was maximal within a few minutes, remained constant for about 1/2 hour and returned to control values over the next 2 to 3 hours. Following return to control, subsequent doses of naloxone produced no further effect on capacity. In chloralose anesthetized animals naloxone also increased the frequency and amplitude of low amplitude pressure waves on the tonus limb of the cystometrogram. Intrathecal administration of naloxone to the sacral spinal cord did not significantly reduce the volume necessary to evoke micturition even at large doses, but did increase the amplitude of micturition contractions. These data, along with previous reports, suggest that mu receptors in the brainstem alter urinary bladder capacity, while delta receptors in the spinal cord modulate the magnitude of bladder contractions. Pharmacological manipulation of these receptor systems could provide a tool for the management of neurogenic bladder dysfunction.

Parasympathetic ganglia: naloxone antagonizes inhibition by leucine-enkephalin and GABA

Synaptic transmission in parasympathetic ganglia of the cat urinary bladder was depressed by low doses (0.1-10 micrograms i.a.) of Leu- or Met-enkephalin but only by larger doses (10 micrograms-1 mg i.a.) of morphine. Naloxone blocked the depressant effects of the opiates as well as the depression produced by GABA, but did not block the depressant effects of norepinephrine. Intracellular recording revealed that Leu-enkephalin reduced EPSP-amplitude and lowered the probability of synaptically evoked firing without altering postsynaptic membrane potential or resistance. These findings suggest that enkephalinergic inhibition in bladder ganglia is mediated at least in part by a presynaptic site of action on delta opiate receptors.

Bladder and urethral sphincter responses evoked by microstimulation of S2 sacral spinal cord in spinal cord intact and chronic spinal cord injured cats.

Urinary bladder and urethral sphincter responses evoked by bladder distention, ventral root stimulation, or microstimulation of S2 segment of the sacral spinal cord were investigated under alpha-chloralose anesthesia in cats with an intact spinal cord and in chronic spinal cord injured (SCI) cats 6-8 weeks after spinal cord transection at T9-T10 spinal segment. Both SCI and normal cats exhibited large amplitude reflex bladder contractions when the bladder was fully distended. SCI cats also exhibited hyperreflexic bladder contractions during filling and detrusor-sphincter dyssynergia during voiding, neither was observed in normal cats. Electrical stimulation of the ventral roots revealed that the S2 sacral spinal cord was the most effective segment for evoking large amplitude bladder contractions or voiding in both types of cats. Microstimulation with a stimulus intensity of 100 microA and duration of 30-60 s via a single microelectrode in the S2 lateral ventral horn or ventral funiculus evoked large amplitude bladder contractions with small urethral contractions in both normal and SCI cats. However, this stimulation evoked incomplete voiding due to either co-activation of the urethral sphincter or detrusor-sphincter dyssynergia. Stimulation in the S2 dorsal horn evoked large amplitude sphincter responses. The effectiveness of spinal cord microstimulation with a single electrode to induce prominent bladder and urethral sphincter responses in SCI animals demonstrates the potential for using microstimulation techniques to modulate lower urinary tract function in patients with neurogenic voiding dysfunctions.