M. Koltzenburg

Activation of unmyelinated afferent fibres by mechanical stimuli and inflammation of the urinary bladder in the cat.

1. We examined the functional properties of unmyelinated primary afferent neurones innervating the pelvic viscera in twenty‐five anaesthetized cats. The axons were isolated from the intact dorsal root and the intact or chronically de‐efferented ventral root of the segment S2. All units were electrically identified with electrical stimulation of the pelvic nerve. 2. The responses of the neurones were studied with natural stimulation of the urinary bladder using innocuous and noxious increases of intravesical pressure and at the onset of an acute artificial inflammation induced by intraluminal injection of mustard or turpentine oil. 3. Out of 297 unmyelinated afferent units isolated from the dorsal root, seven were excited by an increase of the intravesical pressure during contractions and distension of the urinary bladder. These units were silent when the bladder was empty and had thresholds of 30‐50 mmHg which are presumed to be noxious. Further increases of the intravesical pressure were accurately encoded by the discharge rate of the fibres. Out of sixty‐eight unmyelinated afferent units isolated from the ventral root none was activated by these stimuli. 4. Intraluminal injection of mustard oil excited mechanosensitive units at short latency. The discharge was not closely related to changes of the intravesical pressure and the units displayed on‐going activity after the irritant had been removed. This observation suggests that the units had also chemosensitive properties and that the receptive endings were located in the bladder wall. 5. In sixteen cats ninety‐five afferent fibres that were not activated by noxious mechanical stimuli of the urinary bladder were systematically tested with intraluminal injections of mustard oil. This excited 7/67 dorsal root units and 4/28 ventral root units with short latency. Intraluminal application of turpentine oil, tested on twenty‐six afferents in four animals, did not produce a rapid excitation. 6. Following the induction of an inflammation some previously non‐mechanosensitive units started to respond to changes of intravesical pressure in the biologically relevant pressure range of the urinary bladder. 7. In conclusion, a small subpopulation (2.4%) of unmyelinated visceral afferents responds to high, presumably noxious, intravesical pressure and intraluminal application of chemical irritants. Acute inflammation excites a larger proportion of afferents (9.5%) that are not activated by acute noxious mechanical stimulation of the normal urinary bladder. In the inflamed bladder some previously non‐mechanosensitive units started to respond to increases of intravesical pressure. These novel types of chemosensitive receptors may contribute considerably to the pathogenesis of visceral pain states.

Myelinated primary afferents of the sacral spinal cord responding to slow filling and distension of the cat urinary bladder.

1. A total of sixty‐five sacral afferent neurones with myelinated fibres supplying the urinary bladder was recorded from the sacral roots S2 in anaesthetized cats. All afferent units were identified with electrical stimulation of the pelvic nerve. The discharge properties were quantitatively evaluated using slow filling at rates of 1‐2 ml min‐1 and isotonic distension to preset pressure levels. Eight afferents were studied prior to and after acute sacral de‐efferentation of the urinary bladder. 2. All afferent units were silent when the bladder was empty and responded in a graded manner to an increase of intravesical pressure. During slow filling the level of afferent activity correlated closely with the level of the intravesical pressure. All afferents behaved like slowly adapting mechanoreceptors with both a dynamic and static component of their discharge. With the exception of two units the intraluminal pressure threshold was below 25 mmHg. Thus virtually all myelinated afferents respond in the pressure range that is reached during a non‐painful micturition cycle. 3. The stimulus‐response functions of the afferents were similar regardless of whether intravesical pressure was increased by slow filling or by distension. However, during slow filling stimulation response functions often exhibited steeper slopes between 5 and 25 mmHg indicating that relatively small changes of intravesical pressure result in large changes of afferent activity. Nevertheless, all units displayed monotonically increasing stimulus response functions throughout the innocuous and noxious pressure level. 4. The stimulus‐response functions of the afferent neurones did not change after acute de‐efferentation of the urinary bladder, although the rapid phasic fluctuations of afferent activity that are produced by small contractions of the urinary bladder under normal conditions largely disappeared. This means that contractions and distension activate the afferent endings by a common mechanism. 5. It is concluded that the myelinated sacral afferents of the urinary bladder form a homogeneous population which encodes all information necessary for the normal regulation of this organ. Furthermore, this set of afferents mediates all sensations which may reach consciousness within a normal micturition cycle.

Activation of unmyelinated afferents in chronically lesioned nerves by adrenaline and excitation of sympathetic efferents in the cat

Neuroma-in-continuity was experimentally produced in cats by suturing the proximal stump of a transected hindlimb cutaneous nerve to the distal stump of the transected tibial nerve. Eleven to 20 months afterwards single non-myelinated fibres were identified by electrical recording from filaments of the nerves proximally from the neuroma. When tested with repetitive electrical stimulation of the lumbar sympathetic trunk (LST) or i.v. injections of adrenaline, C-fibres in 14 out of 30 filaments were excited by at least one of these stimuli (12 by adrenaline, 7 by stimulation of LST). The fibres responding to LST stimulation were already substantially excited by low frequency stimulation (1-2 Hz). During systemic hypoxia known to result in reflex increases of efferent sympathetic activity responses were elicited in 4 out of 6 C-fibres. Thus, stimuli simulating physiological conditions of sympathetic activity can excite afferent C-fibres in a chronically lesioned nerve.

On the function of spinal primary afferent fibres supplying colon and urinary bladder

The neurophysiological basis of visceral sensations in general and pain in particular have been mainly studied with short-lasting stimuli that simulate the acute events of visceral organ function. Pelvic viscera are supplied by spinal afferents which are involved in the coordinated reflex regulation of continence and evacuation of bowel and bladder and are capable of signalling impending or frank tissue damage. Typically, each afferent neuron innervates one viscus only. The organ-specific subtypes are functionally homogenous and encode by their discharge frequency the information for organ regulation, non-painful and painful sensations. Thus, pain elicited from these organs under physiological conditions is probably not elicited by a specific set of nociceptive visceral afferents. While the use of brief stimuli has yielded invaluable neurophysiological information for normal, healthy viscera, it has fallen somewhat short of providing information about the neuronal basis of chronic visceral pain states. Using pathological models such as experimental inflammation of the urinary bladder and ischaemia of the colon we have shown that the receptor properties of most afferents change dramatically. Of particular interest is the discovery of a novel type of visceral receptor which is not excited by extreme noxious mechanical stimuli applied to the healthy tissue but which is vigorously activated at the onset of an inflammation. This means that the number of functionally active primary afferents is not immutable, but critically depends on the state of the tissue. This new principle of plasticity in the peripheral nervous system bears some considerable importance for the understanding of the genesis of chronic visceral pain states.(ABSTRACT TRUNCATED AT 250 WORDS)

Plasma extravasation in the rat urinary bladder following mechanical, electrical and chemical stimuli: evidence for a new population of chemosensitive primary sensory afferents.

In the skin activation of unmyelinated primary afferents produces vasodilation and plasma extravasation. Here, using Evans blue dye leakage, we have quantitatively compared the ability of mechanical, chemical and electrical stimulation of afferents to produce extravasation in the urinary bladder of anaesthetized rats. Mechanical stimulation - a series of large active contractions of vesical smooth muscle induced by maintained distension - elicited no increase in extravasation compared to controls. In contrast, a similar period of electrical stimulation of nerves supplying the bladder, or chemical stimulation with intravesical instillation of mustard oil, produced high levels of extravasation. We conclude that afferents activated during distension and micturition contractions are different from those which produce extravasation. The latter may be important in pathophysiological cases such as inflammation of the urinary bladder.

A novel type of unmyelinated chemosensitive nociceptor in the acutely inflamed urinary bladder

Single primary afferents supplying the cat urinary bladder were electrophysiologically recorded in the sacral dorsal roots. Under normal conditions, afferents were not spontaneously active when the bladder was empty and innocuous increases of intravesical pressure excited mainly thin myelinated, lowthreshold mechanoreceptors. Less than 2.5% of all unmyelinated visceral afferents responded to a mechanical stimulus, but all had high, presumably noxious thresholds. During an acute inflammation induced with intravesical injections of 2.5% mustard oil or 50–100% turpentine oil both populations developed resting activity and changed their mechanosensitive properties. Out of the many unmyelinated afferents without appreciable mechanosensitivity, an entirely new subpopulation was activated by chemical irritants during an acute inflammation. Subsequently, some of these chemosensitive receptors started to display new mechanosensitive properties. This novel population of sensory neurones may contribute considerably to the pathogenesis of visceral pain states including reflex disturbances of bladder motility.

Dichotomizing unmyelinated afferents supplying pelvic viscera and perineum are rare in the sacral segments of the cat

We have tested the hypothesis that referred pain of pelvic viscera is elicited by dichotomizing branches of unmyelinated primary afferents projecting via the pelvic nerve to the viscera and through the pudendal nerve to the perineum where pelvic pain is commonly referred to. Using neurophysiological techniques 588 unmyelinated single units projecting into either nerve were recorded in the ventral (n = 228) and dorsal (n = 360) root S2. In each sample only one neurone sent an axon into both nerves. Thus, dichotomizing afferents account for less than 0.5% of the afferent neurones and appear to be an unlikely explanation for referred pain in this body area.

Asymmetry and time-course of cutaneous sympathetic reflex responses following sustained excitation of chemosensitive nociceptors in humans

Sympathetic reflex responses were elicited in human volunteers by sustained selective excitation of nociceptors by noxious chemicals, namely topical application of mustard oil which elicited burning pain, or histamine which induced itching in a skin area of 5 cm2 on the volar aspect of one forearm. Stimulus-related sympathetic reflex responses were studied by means of computer-assisted infrared thermography of the palmar aspects of both hands. Nociceptive stimulation induced a decrease of skin surface temperature in both hands interpreted as vasoconstriction. The magnitude of the reflex cooling was correlated with the magnitude of the sensation (r = 0.49), but independent of the quality of sensation (itch or pain). The temperature reduction was maintained for more than 30 min and its time-course matched the time-courses of pain or itch sensations. It is concluded that the sustained and selective excitation of nociceptors elicits a sustained sympathetic reflex response, which adapts very slowly. The time-course of the reflexes suggests that these are not arousal responses, but may be indicators of nociceptive processing in conscious humans. Contralateral temperature decreases were consistently smaller than ipsilateral ones. Thus, sustained nociceptive-specific vasoconstrictor reflexes may be somatotopically organised with an emphasis on areas close to the painful stimulus (homotopic), which has so far only been shown in animals. The study thus demonstrates for the first time in humans the presence of a sympathetic reflex asymmetry, which is specific for nociceptive afferent input.

Plasticity of sympathetic reflex organization following cross-union of inappropriate nerves in the adult cat

1. The present study has investigated the reflex organization of sympathetic neurones and its control of autonomic effector organs following nerve injury and repair. A well‐defined population of vasoconstrictor neurones supplying blood vessels of the hairy skin was forced to innervate a territory that contained some appropriate, but mainly inappropriate autonomic effector organs. For this purpose the central stump of the cut sural nerve was sutured to the peripheral stump of the cut tibial nerve 11‐12 months prior to the terminal experiment. 2. The activity of postganglionic sympathetic neurones was recorded from fine strands of the sural nerve proximal to the nerve lesion. Using a laser‐Doppler device cutaneous blood flow was measured in the hairless skin of the hindpaw that was now reinnervated by the sural nerve. The results show a qualitative change of the reflex organization of sympathetic neurones following cross‐union of these nerves. 3. Stimulation of arterial chemoreceptors by ventilating the animals with a hypoxic gas mixture (8% O2 in N2 for 3‐8 min) increased the activity in twelve out of thirteen strands containing postganglionic sympathetic fibres. The increase of sympathetic activity contrasts with results from normal animals where systemic hypoxia causes a reflex decrease of activity in postganglionic fibres of the sural nerve. 4. Reflex changes of sympathetic activity were closely followed by corresponding changes of cutaneous blood flow. Systemic hypoxia produced vasoconstriction in operated animals in contrast to the vasodilatation observed in normal animals. 5. We conclude that the reflex organization of sympathetic neurones can change qualitatively following nerve lesion when sympathetic neurones regenerate and supply inappropriate target tissues. This long‐lasting change reflects the plasticity of the autonomic nervous system and can produce a sustained abnormal control of reinnervated autonomic effector organs.

RECEPTIVE PROPERTIES OF PIAL AFFERENTS

&NA; The blood vessels and the pial surface of the brain, spinal cord and its roots are innervated by primary afferent neurones. Here, we have electrophysiologically characterized the functional properties of a subpopulation of these afferent fibres that supply the ventral roots of the cat sacral spinal cord. We have taken advantage of the unique anatomical arrangement of these primary afferent neurones which have their central axon in the dorsal root and project with their peripheral process into the segmental ventral root. In 10 experiments, 14 units were recorded in the dorsal root S2 which responded to electrical stimulation of the segmental ventral root. As judged by their conduction velocity ranging from 0.1 to 2.3 m/sec, all fibres were unmyelinated. In 4 cases a spot‐like receptive field was located on the root where the units were reproducibly activated by mechanical stimuli, the most effective being a slight stretch. In two units tested, topical application of hypertonic saline onto the receptive field, but not at other portions of the axon elicited a long‐lasting vigorous discharge with intermittent bursts. There was no obvious association of the receptive field with small blood vessels. In 5 of the 14 units including 2 with a mechanosensitive receptive field we observed latency jumps of the action potential with electrical stimulation of the ventral root close to the dorsal root ganglion. In some of these units latency jumps were also observed at other positions when the stimulation electrodes were moved centrally towards the spinal cord. We conclude that a subpopulation of unmyelinated fibres in the spinal ventral root are primary afferents innervating the root proper or its sheath. Since the fibres responded best to noxious stimuli they are a potential peripheral pathway involved in a number of radicular pain states. The presence of latency jumps suggests that some of these afferents have multiple axon collaterals.