David F. Cottrell

Analgesia mediated by the TRPM8 cold receptor in chronic neuropathic pain.

BACKGROUND Chronic established pain, especially that following nerve injury, is difficult to treat and represents a largely unmet therapeutic need. New insights are urgently required, and we reasoned that endogenous processes such as cooling-induced analgesia may point the way to novel strategies for intervention. Molecular receptors for cooling have been identified in sensory nerves, and we demonstrate here how activation of one of these, TRPM8, produces profound, mechanistically novel analgesia in chronic pain states. RESULTS We show that activation of TRPM8 in a subpopulation of sensory afferents (by either cutaneous or intrathecal application of specific pharmacological agents or by modest cooling) elicits analgesia in neuropathic and other chronic pain models in rats, thereby inhibiting the characteristic sensitization of dorsal-horn neurons and behavioral-reflex facilitation. TRPM8 expression was increased in a subset of sensory neurons after nerve injury. The essential role of TRPM8 in suppression of sensitized pain responses was corroborated by specific knockdown of its expression after intrathecal application of an antisense oligonucleotide. We further show that the analgesic effect of TRPM8 activation is centrally mediated and relies on Group II/III metabotropic glutamate receptors (mGluRs), but not opioid receptors. We propose a scheme in which Group II/III mGluRs would respond to glutamate released from TRPM8-containing afferents to exert an inhibitory gate control over nociceptive inputs. CONCLUSIONS TRPM8 and its central downstream mediators, as elements of endogenous-cooling-induced analgesia, represent a novel analgesic axis that can be exploited in chronic sensitized pain states.

Neurofascins Are Required to Establish Axonal Domains for Saltatory Conduction

Voltage-gated sodium channels are concentrated in myelinated nerves at the nodes of Ranvier flanked by paranodal axoglial junctions. Establishment of these essential nodal and paranodal domains is determined by myelin-forming glia, but the mechanisms are not clear. Here, we show that two isoforms of Neurofascin, Nfasc155 in glia and Nfasc186 in neurons, are required for the assembly of these specialized domains. In Neurofascin-null mice, neither paranodal adhesion junctions nor nodal complexes are formed. Transgenic expression of Nfasc155 in the myelinating glia of Nfasc-/- nerves rescues the axoglial adhesion complex by recruiting the axonal proteins Caspr and Contactin to the paranodes. However, in the absence of Nfasc186, sodium channels remain diffusely distributed along the axon. Our study shows that the two major Neurofascins play essential roles in assembling the nodal and paranodal domains of myelinated axons; therefore, they are essential for the transition to saltatory conduction in developing vertebrate nerves.

Restricted growth of Schwann cells lacking Cajal bands slows conduction in myelinated nerves

Nerve impulses are propagated at nodes of Ranvier in the myelinated nerves of vertebrates. Internodal distances have been proposed to affect the velocity of nerve impulse conduction; however, direct evidence is lacking, and the cellular mechanisms that might regulate the length of the myelinated segments are unknown. Ramón y Cajal described longitudinal and transverse bands of cytoplasm or trabeculae in internodal Schwann cells and suggested that they had a nutritive function. Here we show that internodal growth in wild-type nerves is precisely matched to nerve extension, but disruption of the cytoplasmic bands in Periaxin-null mice impairs Schwann cell elongation during nerve growth. By contrast, myelination proceeds normally. The capacity of wild-type and mutant Schwann cells to elongate is cell-autonomous, indicating that passive stretching can account for the lengthening of the internode during limb growth. As predicted on theoretical grounds, decreased internodal distances strikingly decrease conduction velocities and so affect motor function. We propose that microtubule-based transport in the longitudinal bands of Cajal permits internodal Schwann cells to lengthen in response to axonal growth, thus ensuring rapid nerve impulse transmission.

Peripheral demyelination and neuropathic pain behavior in periaxin-deficient mice

The Prx gene in Schwann cells encodes L- and S-periaxin, two abundant PDZ domain proteins thought to have a role in the stabilization of myelin in the peripheral nervous system (PNS). Mice lacking a functional Prx gene assemble compact PNS myelin. However, the sheath is unstable, leading to demyelination and reflex behaviors that are associated with the painful conditions caused by peripheral nerve damage. Older Prx-/- animals display extensive peripheral demyelination and a severe clinical phenotype with mechanical allodynia and thermal hyperalgesia, which can be reversed by intrathecal administration of a selective NMDA receptor antagonist We conclude that the periaxins play an essential role in stabilizing the Schwann cell-axon unit and that the periaxin-deficient mouse will be an important model for studying neuropathic pain in late onset demyelinating disease.

Afferent activity in the superior spermatic nerve of lambs — The effects of application of rubber castration rings

Electrophysiological techniques were used to record afferent activity in the superior spermatic nerves of young lambs under general anaesthesia. Receptive fields were identified in the pampiniform plexus and the deep tissue of the testis in response to mechanical stimulation. Application of a standard rubber castration ring to the scrotal neck evoked vigorous afferent activity, including some from formerly silent units with receptive fields particularly in the pampiniform plexus. Some of this multi-unit discharge adapted rapidly within 10 s of the application of the ring and was followed by a discharge pattern which decayed exponentially over 90 min. The rate of decay of this discharge showed more than one exponent (time constant) with inflections at approximately 90 s and 16 min. After the application of the castration ring, quantitatively controlled scrotal compression continued to excite receptors, though a declining frequency was recorded over the period of observation. It was concluded that: (a) rubber castration rings initiated afferent activity which persisted for periods in excess of 90 min, a time course which is similar to the behavioural and humoral changes in the conscious animal; (b) both standard and small rubber castration rings were ineffective in rapidly producing neuronal pressure block of the slowly conducting afferent fibres in the superior spermatic nerve; (c) intra-testicular injection of local anaesthetic rapidly blocked afferent fibres running in the superior spermatic nerve.

Effect of Limb Lengthening on Internodal Length and Conduction Velocity of Peripheral Nerve

The influences of axon diameter, myelin thickness, and internodal length on the velocity of conduction of peripheral nerve action potentials are unclear. Previous studies have demonstrated a strong dependence of conduction velocity on internodal length. However, a theoretical analysis has suggested that this relationship may be lost above a nodal separation of ∼0.6 mm. Here we measured nerve conduction velocities in a rabbit model of limb lengthening that produced compensatory increases in peripheral nerve growth. Divided tibial bones in one hindlimb were gradually lengthened at 0.7 mm per day using an external frame attached to the bone. This was associated with a significant increase (33%) of internodal length (0.95–1.3 mm) in axons of the tibial nerve that varied in proportion to the mechanical strain in the nerve of the lengthened limb. Axonal diameter, myelin thickness, and g-ratios were not significantly altered by limb lengthening. Despite the substantial increase in internodal length, no significant change was detected in conduction velocity (∼43 m/s) measured either in vivo or in isolated tibial nerves. The results demonstrate that the internode remains plastic in the adult but that increases in internodal length of myelinated adult nerve axons do not result in either deficiency or proportionate increases in their conduction velocity and support the view that the internodal lengths of nerves reach a plateau beyond which their conduction velocities are no longer sensitive to increases in internodal length.

1 – Regulation of Gut Motility by Luminal Stimuli in the Ruminant

Publisher Summary Luminal stimuli play an important role in the regulation of gastrointestinal motility. The volume and composition of digesta initiate motility patterns by activating neural and hormonal mechanisms. The chapter deals with the neural mechanisms involved and concentrates on experiments utilizing physiological preparations where the stimuli are within normal limits. Using this approach makes it easy to deduce the importance of the mechanisms involved in the intact animal. Luminal stimuli include distention, tactile, chemical, viscosity, and possibly thermal changes. The regulatory mechanisms by luminal stimuli are modified by factors such as fasting, inherent circadian rhythms, and the postprandial release of GI-tract and other hormones—all of which require further study and appropriate experimental procedures.

Light Microscopy of the Enteric Nervous System of Horses with or without Equine Dysautonomia (Grass Sickness): its Correlation with the Motor Effects of Physostigmine

Light microscopy was undertaken on sections from the caudal flexure of the duodenum and the terminal ileum proximal to the ileocaecal fold in 5 control horses, 5 horses with acute grass sickness (AGS), and 5 horses with chronic grass sickness (CGS). With the exception of the ileal submucous plexus of the CGS group, the AGS group had the lowest number of neurons as measured using a subjective scoring scheme. The proportion of abnormal neurons in the AGS group was similar in both plexuses and both regions, whereas the values for the CGS group were much higher in the duodenal region than in the ileal region. The motility of tissue adjacent to that used for histology was measured isometrically in vitro. The increase in the rate of contractions following exposure to physostigmine was greatest for the AGS group, both from the duodenal and from the ileal region. The latency was longest for the AGS group, suggesting that the material from this group had the least number of active cholinergic neurons. The studies with physostigmine thus indicated that the most severe functional damage occurred in cases of AGS. These findings confirm that extensive damage occurs in the enteric neurons in equine grass sickness. There was good correlation between the functional cholinergic responses and the extent of neuronal degeneration.

Vagal reflex inhibition of motility in the abomasal body of sheep by antral and duodenal tension receptors

Vagally-mediated regulation of motility in the abomasal body by duodenal and abomasal antral motility was demonstrated in acutely prepared anaesthetized sheep. The enteric plexuses between the abomasal body, antrum and duodenum were interrupted by transection. Antral contractions were more effective than duodenal contractions at causing inhibition of the abomasal body, and antral isometric conditions were more effective than antral isotonic conditions. Inhibition of motility in the abomasal body was reduced by unilateral cervical vagotomy, was abolished by bilateral cervical vagotomy, and was reversibly inhibited by cervical vagus cold block. The demonstration of vagal pathways in abomaso-abomasal reflexes confirms a functional homology of the mechanisms in the ruminant and animals with simple forms of stomach.

A mechanical stimulator for the determination of the minimum alveolar concentration (MAC) of halothane in the rabbit

The minimum alveolar concentration (MAC) of halothane was determined in New Zealand White rabbits. Tracheal anaesthetic concentrations were measured using a Siemens Servo Gas Monitor. A stimulator was used to deliver precisely controlled mechanical stimuli for the determination of MAC. Movement of the rabbits head was recorded using a force transducer attached to the teeth. Evidence is presented that for the determination of MAC a precise nociceptive threshold is preferable to the so-called supramaximal stimulus used in clinical anaesthesia and in determinations of anaesthetic potency. We conclude that techniques for the determination of MAC which disregard either sensitization of sensory mechanisms by producing tissue inflammation or the possibility of nerve compression by severe mechanical stimuli are of questionable value. The use of the mechanical stimulator described, or a similar device, would help in the standardization of the determination of MAC in all species by facilitating the application of a force of controlled amplitude, duration and velocity, thereby removing some of the variables which confound comparative studies of MAC.