Ryohji Nakamura

Late-onset selective neuronal damage in the rat spinal cord induced by continuous intrathecal administration of AMPA

Neurotoxicity mediated by 1-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) was investigated by infusing this agent continuously for 7 days intrathecally to adult rats using a mini-osmotic pump. Behavioral changes were apparent only after the second postoperative day, when the rats displayed hindlimb palsy or incontinence of urine. The behavioral deficits became progressively severe and the rats usually displayed both hindlimb paraplegia and incontinence of urine by the 7th postoperative day. These progressive behavioral deficits were induced in a dose-dependent manner in the rats that received AMPA at a dose of > 100 pmol/h (100 microM at 1 microliter/h, 17 nmol in total dose). The severity of behavioral deficits was in parallel with that of neuropathological changes in the lumbosacral cords. In spinal segments rostrally adjacent to those with severe pathological changes, only the neurons in the dorsal horns (Rexeds laminae II-IV) were destroyed with intense gliosis. These changes were not induced by infusing AMPA for 1 day. The concomitant administration of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), an antagonist for non-N-methyl-D-aspartate (NMDA) receptors, with AMPA, but not that of 2-amino-5-phosphonovalerate (APV), an antagonist for NMDA receptor, prevented induction of the behavioral and neuropathological changes. The findings of the present study suggest that this late-onset, selective neurotoxicity is mediated by AMPA-type glutamate receptors.

AMPA receptor-mediated slow neuronal death in the rat spinal cord induced by long-term blockade of glutamate transporters with THA

Excitotoxicity secondary to the loss of glutamate transporters (GluT) has been proposed as a possible pathogenetic mechanism for neuronal degeneration in amyotrophic lateral sclerosis. We therefore investigated whether prolonged in vivo pharmacologic inhibition of GluT would result in neuronal damage in the rat. DL-Threo-beta-hydroxyaspartate (THA), a potent GluT inhibitor, and glutamate were continuously infused into the rat spinal subarachnoid space by using a mini-osmotic pump. Animals that received both THA and glutamate, but not those received either singly, displayed tail paralysis with or without hind-limb paralysis and urinary incontinence after the third postoperative day. Pathologically, symptomatic animals exhibited neuronal loss with a variable extent of gliosis preferentially involving the dorsal horn of the lumbosacral cord. In the rostral spinal segments adjacent to those regions of intense pathologic changes, small neurons in the dorsal horn were selectively destroyed, a pattern similar to the late-onset neuronal damage induced by continuous intrathecal administration of 1-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) [R. Nakamura et al., Brain Res. 654 (1994) 279-285]. These behavioral and pathologic changes were blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), suggesting that pharmacologic blockade of GluT causes selective neuronal damage in vivo by AMPA receptor activation.

Somatosensory Conduction Delay in Central and Peripheral Nervous System of Diabetic Patients

Fifty-four diabetic patients with or without clinical evidence of neuropathy and with no clinical evidence of CNS dysfunction were studied by somatosensory-evoked potentials after electrical stimulation of the median nerve at the wrist and recorded from the scalp electrode against a noncephalic reference. Peripheral conduction index, calculated as the distance from the wrist to the C7 spinous process divided by the P9 latency, was significantly decreased (P <0.01) in diabetic patients (69.81 ± 6.47 m/s) compared with 28 age-matched nondiabetic subjects (76.85 ± 5.65 m/s). The P11–P13 interpeak latency, representative of the transit time from the dorsal column at the level of the sensory input into the cervical cord to the brain stem along the somatosensory pathways (CCT1), and the P13-N19 interpeak latency, representative of the transit time from the brain stem to the somatosensory cortex (CCT2), were significantly increased in diabetic patients (CCT1, 2.51 ± 0.63 ms; CCT2, 5.76 ± 0.92 ms) compared with nondiabetic subjects (CCT1, 2.28 ± 0.36 ms, P <0.05; CCT2, 5.18 ± 0.51 ms, P <0.01). We conclude that, in diabetic patients, neurophysiological abnormalities may be present in two distinct parts of the CNS and the peripheral nervous system.

Harmful effects of anti-GalNAc-GD1a antibodies and TNF-alpha on rat dorsal root ganglia.

Abstract  The clinical characteristics of five (22%) of 23 patients with Guillain‐Barré syndrome (GBS), whose serum contained immunoglobulin G (IgG) antibodies to the ganglioside N‐acetylgalactosaminyl GD1a (GalNAc‐GD1a), included pure motor weakness of the axonal type. These patients had a relatively good prognosis, but displayed higher serum tumor necrosis factor‐alpha (TNF‐α) titers than the other GBS patients. We examined the effect of serum from these patients with IgG anti‐GalNAc‐GD1a antibodies on neurites from cultured rat dorsal root ganglia (DRG) and found it to damage the myelin in well‐elongated DRG neurites and monolayer cultures of Schwann cells and neurons. In the regeneration model, serum from these patients delayed neurite extension and inhibited Schwann cell proliferation. Neurons in cultured monolayers showed vacuolation and decreased rapidly in number. Schwann cells were also vacuolated and readily detached from the substratum. The effects of IgG anti‐GalNAc‐GD1a antibodies purified from one of the patients, rabbit serum after immunization with GalNAc‐GD1a, and recombinant TNF‐α were also examined. IgG anti‐GalNAc‐GD1a antibodies mainly inhibited the regeneration and preservation of neurons, while TNF‐α mainly induced morphological changes in well‐proliferated Schwann cells and myelin.

Concentration-dependent changes in motor behavior produced by continuous intrathecal infusion of excitatory amino acids in the rat spinal cord

A growing body of evidence has suggested that glutamate receptors mediate selective degeneration of neurons in the central nervous system during the development of neurodegenerative diseases. Glutamate receptors are divided into N-methyl-D-aspartate (NMDA)-type and non-NMDA-type. Neurotoxicity mediated by the latter has attracted much interest as a possible causative mechanism underlying amyotrophic lateral sclerosis (ALS). As the clinical course of ALS is chronic and progressive, investigation of chronic effects of non-NMDA receptor agonists on neuronal function would be useful for evaluating the role of glutamate receptor-mediated neurotoxicity in ALS. However, chronic non-NMDA receptor-mediated neurotoxicity has been investigated less thoroughly than acute neurotoxicity. We infused an aqueous solution of R,S-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) intrathecally and continuously by an osmotic minipump in rats. This method of continuous infusion enabled us to keep the drug concentration relatively constant in the cerebrospinal fluid surrounding the spinal cord. The present method of AMPA administration is more suitable for investigating ALS pathogenesis than acute injections, in view of the gradual progression of the disease and the selectivity of lesions produced.