PhD

dissertationThe dose-response effects of clonidine on transmission through somatospinal reflex, viscerospinal reflex, intraspinal, and spinal-bulbospinal reflex pathways were determined in spinal or chloralose-anesthetized cats to assess principal sites of drug action. Clonidine rapidly produced parallel, dose-dependent depression of transmission through each pathway which was antagonized by tolazoline or yohimbine. The order of descending sensitivity was found to be spinal-bulbospinal, intraspinal, and spinal reflex pathway. Analysis of the relative depression of transmission at spinal and at brainstem levels indicates that the spinal site is more sensitive to clonidine that it is generally considered to be. The effect of the serotonin (5-HT) precursor 5-hydroxytryptophan (5-HTP) was also assessed on the intraspinal pathway and somatospinal reflex. In contrast to clonidine, 5-HTP was more effective in depressing the spinal reflex than the intraspinal pathway, and both pathways could be depressed completely. Clonidine and 5-HTP appear to depress the excitability of sympathetic neurons by activating alpha(,2)- and 5-HT receptors, respectively. The intraspinal pathway was rapidly and markedly enhanced for 1-2 hours by two methylxanthines. Clonidine depressed intraspinal transmission and prevented enhancement by the xanthines; alpha(,2)-receptor antagonists blocked the effect of clonidine and not only restored but also markedly prolonged the enhancement by the xanthines. The results suggest that the excitability of sympathetic preganglionic neurons may be regulated by cyclic AMP through activation of different subtypes of adrenergic receptors that are either positively or negatively coupled to adenylate cyclase. Methyldopa (MD) produced a moderate enhancement of transmission through three central sympathetic pathways three hours after an i.v. infusion of 150 mg/kg. However, a subsequent dose of 5 mg/kg dose of reserpine, which alone causes no depression, produces prompt, marked depression of transmission through each pathway which is antagonized by yohimbine, suggesting that reserpine releases an active metabolite of MD to depress sympathetic preganglionic neurons by activing alpha(,2)-receptors. Depletion of epinephrine by blockade of phenylethanolamine-N-methyltransferase prevents this depression that occurs with the transmitter release. Propranolol modestly enhances transmission through the pathways tested

A Guidance Channel Seeded With Autologous Schwann Cells for Repair of Cauda Equina Injury in a Primate Model

Background/Objective: To evaluate an implantable guidance channel (GC) seeded with autologous Schwann cells to promote regeneration of transected spinal nerve root axons in a primate model. Methods: Schwann cells were obtained from sural nerve segments of monkeys (Macaca fascicularis; cynomolgus). Cells were cultured, purified, and seeded into a PAN/PVC GC. Approximately 3 weeks later, monkeys underwent laminectomy and dural opening. Nerve roots of the L4 through L7 segments were identified visually. The threshold voltage needed to elicit hindlimb muscle electromyography (EMG) after stimulation of intact nerve roots was determined. Segments of 2 or 3 nerve roots (each ∼8-15 mm in length) were excised. The GC containing Schwann cells was implanted between the proximal and distal stumps of these nerve roots and attached to the stumps with suture. Follow-up evaluation was conducted on 3 animals, with survival times of 9 to 14 months. Results: Upon reexposure of the implant site, subdural nerve root adhesions were noted in all 3 animals. Several of the implanted GC had collapsed and were characterized by thin strands of connective tissue attached to either end. In contrast, 3 of the 8 implanted GC were intact and had white, glossy cables entering and exiting the conduits. Electrical stimulation of the tissue cable in each of these 3 cases led to low-threshold evoked EMG responses, suggesting that muscles had been reinnervated by axons regenerating through the repair site and into the distal nerve stump. During harvesting of the GC implant, sharp transection led to spontaneous EMG in the same 3 roots showing a low threshold to electrical stimulation, whereas no EMG was seen when harvesting nerve roots with high thresholds to elicit EMG. Histology confirmed large numbers of myelinated axons at the midpoint of 2 GC judged to have reinnervated target muscles. Conclusions: We found a modest rate of successful regeneration and muscle reinnervation after treatment of nerve root transection with a Schwann cell-seeded, implanted synthetic GC. Newer treatments, which include the use of absorbable polymers, neurotrophins, and antiscar agents, may further improve spinal nerve regeneration for repair of cauda equina injury

Innervation and Properties of the Rat FDSBQ Muscle: An Animal Model to Evaluate Voluntary Muscle Strength after Incomplete Spinal Cord Injury

Muscles innervated from spinal segments close to the site of a human spinal cord injury are often under voluntary control but are weak because they are partially paralyzed and partially denervated. Our objective was to develop an animal model of this clinical condition to evaluate strategies to improve voluntary muscle strength. To do so, we examined the spinal and peripheral innervation of the flexor digitorum superficialis brevis quinti (FDSBQ) muscle of the rat foot, characterized the muscle and motor unit properties, and located the FDSBQ motoneurons. Retrograde labeled motoneurons were in L4 to L6 spinal cord. Unilateral stimulation of L4 to S1 ventral roots and recording of evoked force showed that FDSBQ motor axons exited via two ventral roots (L5 and L6 or L6 and S1) in 38% of rats and via one ventral root in 62% of rats. FDSBQ motor axons traveled via two peripheral nerves, the lateral plantar (76% of axons) and sural nerves (24%). Each ventral root contributed motor axons to each nerve branch. Thus, by combining conduction block of one peripheral nerve to induce partial muscle paralysis and ventral root section to induce partial denervation, it is possible to produce in one rat muscle the consequences of many human cervical spinal cord injuries. FDSBQ muscles and motor units were mainly fast-twitch, fatigable, and composed of fast-type muscle fibers. The narrow range of motor unit forces (1–13 mN), the low mean twitch force (5.1±0.3 mN), and the large number of motoneurons (31±4) suggest that rat FDSBQ muscle is a good model of distal human musculature which is frequently influenced by spinal cord injury. We conclude that the FDSBQ muscle and its innervation provide a useful animal model in which to study the consequences of many spinal cord injuries which spare some descending inputs but also induce substantial motoneuron death near the lesion