Noncontusive segmental spinal cord injury using radiofrequency current.

OBJECTIVE: To develop a nondisruptive model for the study of spinal cord injury. METHODS: A 2-MHz radiofrequency heating chamber was mounted over the rat T13-L1 vertebral column via a short dorsal incision. Epidural temperature at chamber level was monitored via a small proximal laminotomy. Forty-three rats were studied using time-temperature heating regimens from 45 to 48.5 degrees C and 4 to 15 minutes. A blinded numerical hind limb impairment score (Neurologic Impairment Score) was determined at intervals up to 2 weeks after injury. Segmental spinal cord blood flow was measured using [14C]butanol tissue uptake in injured and control rats. RESULTS: Above the injury threshold, increasing the time-temperature regimens was associated with a progressively worse Neurologic Impairment Score (r = 0.73-0.87 up to 24 hours after injury). Cord blood flow was unchanged at 2 hours but was 44% depressed at the injury level 6 hours after injury (p < 0.01). Histologically, injury extended minimally beyond the injured segment. Vascular thrombosis was not seen. CONCLUSION: This comparatively noninvasive model does not mechanically disrupt cord components and results in progressive neurologic impairment that correlates with the time-temperature regimen used for injury. It should be useful in identifying secondary phenomena that worsen functional status after cord trauma.

Methylprednisolone treatment of experimental spinal cord injury.

UNLABELLED: Lipid peroxidation has been identified as a deleterious consequence of contusive spinal cord trauma and of thermal injury. The L3-L6 spinal cord segment was thermally injured using a radiofrequency heating chamber mounted on the vertebral column of anesthetized rats. Hind limb function was assessed 2 hours later. A bolus of methylprednisolone (MP, 30 mg/kg) was then given intravenously, followed by 5.4 mg/kg/hr MP for 6 or 24 hours. Cord water content and regional spinal cord blood flow (RSCBF, 14C-butanol distribution) were measured at seven cord levels after function had been reassessed following treatment. Untreated rats were given vehicle. The study was randomized and blinded. RESULTS: Edema in heated segments was progressive over 24 hours, but was the same in treated vs untreated rats. RSCBF in heated segments was the same in treated vs uninjured controls at 6 and 24 hours. In untreated rats, RSCBF in the heated segment was elevated by 30% at 6 hours, but was the same as uninjured control by 24 hours. In the unheated segments of untreated rats, RSCBF was elevated at 24 hours. At 24 hours, RSCBF was lower in treated vs untreated rats at all levels, including the heated one. Limb function deteriorated equivalently in both groups. CONCLUSION: MP obviated the early rise in RSCBF in heated segments and the elevations in RSCBF in uninjured segments, but had no effect on cord edema or on limb function.

Effects of 4-aminopyridine on protein phosphorylation in heat-blocked peripheral nerve.

The topical application of 4-aminopyridine (4-AP) reverses within 1-3 min the partial conduction block that results from heating 5-mm segments of rat posterior tibial, peroneal and sural nerves to 45 degrees C for several minutes. Nerves previously blocked in vitro or in vivo were incubated in vitro in the presence of [gamma-32P]ATP. The rate of phosphorylation of soluble nerve proteins that had entered the media was determined. Labeled proteins were separated electrophoretically and autoradiograms prepared. We found that 5 mM 4-AP increases the phosphorylation rate in heat-blocked nerve by approximately 50-fold. The process is calcium-dependent and is heat-labile. Soluble proteins with a molecular weight in the 53-55 kDa range are preferentially but not exclusively phosphorylated in the presence of 4-AP at levels effective in restoring conduction. The results suggest that the potassium channel blocker 4-AP may restore conduction by inducing changes in channel proteins.

Effect of adrenalectomy or sympathectomy on spinal cord blood flow in hypothermic rats.

We used [14C]butanol distribution to measure regional spinal cord blood flow (RSCBF) in three groups of pentobarbital-anesthetized rats: 1) normals (n = 16); 2) after bilateral adrenalectomy (n = 18); and 3) after excision of the abdominal sympathetic chains (n = 12). Half of the rats in each group were maintained near normothermia; the remainder were colled to a rectal temperature of 25-27 degrees C. In intact rats, there was a 24-37% increase in RSCBF during hypothermia in the cord region C3-L6 inclusive. Neither operative procedure altered RSCBF during normothermia. In hypothermic adrenalectomized rats, RSCBF was elevated only in the C3-C5 cord segment. After sympathectomy, RSCBF was unchanged during hypothermia. In the cauda equina, flow fell in all hypothermic rats. The hypothermia-associated increases in RSCBF were not related to changes in mean arterial blood pressure. We conclude that adrenalectomy near-totally ablates the hypothermia-associated increase in RSCBF measured in intact rats and that abdominal sympathectomy totally ablates it. This evidence complements morphological evidence for adrenergic innervation of the spinal cord vasculature.

4-aminopyridine restores conduction in heat-blocked sciatic branches.

We induced partial conduction block in rat sural and posterior tibial nerves by heating a short (5 mm) segment of the midportions of these branches to 45 degrees C until the amplitude of the elicited compound action potential was reduced by at least 50%, an effect that appeared to be irreversible without specific treatment. Excised branches studied in vitro were compared with branches heated in situ with an intact blood supply. The potassium channel blocker 4-aminopyridine (4-AP, 5 mM) was applied topically to the heated segments at intervals after injury. The amplitude of the compound action potential was restored to the control level in all preparations treated within 10 minutes after the induction of conduction block. The reversal appeared to be more rapid in the in vivo preparations, and it persisted for at least 60 minutes. The data suggest that paranodal potassium channel exposure may accompany direct thermal injury to the peripheral nerve.

Regional blood flow in resting and stimulated sciatic nerve of diabetic rats.

The role of ischemia in the pathogenesis of diabetic peripheral neuropathy remains uncertain. We used the distribution of [14C]butanol to measure resting regional sciatic nerve blood flow in normal, anesthetized rats and in rats with acute experimental diabetes from streptozotocin administration. Regional flows in hind limb biceps femoris muscle and skin were simultaneously measured. In additional diabetic rats, these blood flows were compared in both limbs after proximal electrical stimulation of one sciatic trunk (10 shocks/s) for 15 min. One month after streptozotocin administration, 8 of 11 test rats were hyperglycemic. Resting nerve blood flow in the hyperglycemic rats--5.6 +/- 3.07 ml.min-1.100 g-1--was significantly less than that in the controls (9.4 +/- 3.9 ml.min-1.100 g-1, P = 0.002). Muscle blood flow was normal and skin blood flow decreased in these rats. Calculated tissue vascular resistances were elevated in all three tissues. Stimulation of one sciatic trunk in five other diabetic rats resulted in a stimulated nerve blood flow of 15.7 +/- 7.7 ml.min-1.100 g-1, and nerve blood flow in the resting control limb was 7.7 +/- 4.3 ml.min-1.100 g-1 (P = 0.009). Muscle blood flow increased approximately fourfold on the stimulated side but skin blood flow did not increase. Resting sciatic nerve blood flow is modestly decreased in acute streptozotocin-induced diabetes, but the neural blood vessels are still responsive to the increase in nerve metabolic activity associated with nerve stimulation.

Sciatic nerve function following hindlimb thermal injury.

Peripheral neuropathy occurs in approximately 20% of patients with major burns and seriously impairs rehabilitation. We describe an experimental model which permits elevation of the tissue temperature in the region of the distal sciatic nerve trunk of rats at a reproducible rate to a predetermined level without inflicting concomitant major cutaneous injury. Radiofrequency current is delivered through parallel copper electrodes mounted in a chamber into which the limb has been inserted. In the present experiments, tissue temperature was arbitrarily elevated to 47 degrees C for 30 sec in 62 rats. There were 43 normal controls. The posterior tibial branch was the most intensively studied, as some of its conduction characteristics can be serially assessed percutaneously. Conduction block, which was apparently irreversible, was present in 67% of posterior tibials by 24 hr postinjury. In branches which were still excitable, prolongation of the absolute refractory period was the most consistent abnormality noted. Slowing of conduction, as evidenced by prolongation of inflection velocity or peak velocity, was never observed. However, this injury resulted in selective conduction failure of sural--but not of peroneal--fibers which conducted at 40 m/sec or greater. Fiber modality is an important determinant of the vulnerability to direct thermal injury of peripheral nerve in vivo.

Regional sciatic nerve and muscle blood flow in conscious and anesthetized rats.

The regulation of peripheral nerve blood flow is incompletely understood. Regional blood flow in the rat sciatic nerve (NBF) and hamstring muscle (MBF) was measured in both conscious and anesthetized normal rats and in rats that had undergone surgical exposure of one sciatic nerve just prior to measurement. The distribution of [14C]butanol following its bolus intravenous injection was used to determine the flows in a modification of the "indicator fractionation" technique. NBF in normal rats was similar in limb pairs and was unaffected by pentobarbital sodium anesthesia. The pooled value was 12.5 +/- 1.1 ml X min-1 X 100 g-1. NBF was unaffected by sham operation in the conscious rats but was doubled in operated limbs of anesthetized rats (P less than 0.001). MBF in conscious normal rats was five times that measured during anesthesia. As in NBF, sham operation significantly increased MBF only in anesthetized rats (P less than 0.01). [14C]butanol distribution is a sensitive indicator of NBF and MBF. Mere surgical exposure of the nerve significantly increases NBF and MBF in anesthetized, but not in conscious rats.

Potassium ion channel blockade restores conduction in heat-injured nerve and spinal nerve roots.

Compound action potentials were recorded in vitro from rat peroneal and sural nerves and from dorsal and ventral roots of the cauda equina before and after radiofrequency heating of 5-mm-length segments of these nerves to 41 to 45 degrees C. The heating was continued for intervals sufficient to reduce response amplitude by 50%. Inflection velocity, potential duration at 1/2 peak height, and the proportion of conducting A alpha fibers were also measured. The topical application of 4-aminopyridine (4-AP) and tetraethylammonium chloride (TEA) to the previously heated segments immediately following the radiofrequency injury completely or near-completely restored amplitude height to the preheat value in all experiments. A alpha sensory fibers were the most susceptible to the conduction block. Conduction in these fibers was also the most readily restored by the application of 4-AP or TEA. The effects of TEA, but not of 4-AP, could be reversed by saline or buffer washing. Topical application of verapamil and of magnesium or calcium ions had no discernible effect on heated nerves. We suggest that the mechanism of heat-induced conduction block may be similar to that from early demyelination or stretch injury. Further, motor and sensory A alpha fibers differ both in their vulnerability to heat and in their subsequent response to the application of potassium channel blockers.

Differential effects of in vitro heating on rat sciatic nerve branches and spinal nerve roots.

Locally applied heat induces nerve conduction block. Conflicting observations have been made regarding the relation of fiber conduction velocity to heat sensitivity. This study utilized sciatic nerve branches and spinal nerve roots which were heated until a substantial conduction block occurred. The results indicated that sensory fibers conducting at greater than 40 m/s are more heat-sensitive than motor fibers of the same conduction velocity.

Stimulation of thiamine triphosphate metabolism in nerve by cyclic guanosine 3',5'-monophosphate.

This paper reports that addition of cGMP results in an increase of the amount of(32)Pi incorporated into thiamine triphosphate in nerve roots and sympathetic trunks. The effect is present both at rest and during electrical stimulation. Other nucleotides were less effective. Theophylline increased the incorporation, possibly because of phosphodiesterase inhibition. A blocking effect of atropine was noted in sympathetic trunk preparations that contain ganglion cells.

Correlation between gait speed and spasticity at the knee.

An automated system designed and developed at this laboratory was used to measure spasticity in the thigh musculature of fifteen patients with hemiplegia. In addition, each subject was timed during a gait trial over a measured distance at maximum speed. Analysis of the data revealed no statistically significant correlation between spasticity in the thigh musculature and the gait speed of the subjects studied. Factors which may have contributed to the findings are mentioned.