Using transcranial magnetic stimulation (TMS), we investigated how short-interval intracortical inhibition (SICI) was involved with transient motor cortex (M1) excitability changes observed just before the transition from muscle contraction to muscle relaxation. Ten healthy participants performed a simultaneous relaxation task of the ipsilateral finger and foot, relaxing from 10% of their maximal voluntary contraction (MVC) force after the go signal. In the simple reaction time (RT) paradigm, single or paired TMS pulses were randomly delivered after the go signal, and motor evoked potentials (MEPs) were recorded from the right first dorsal interosseous (FDI) muscle. We analyzed the time course prior to the estimated relaxation reaction time (RRT), defined here as the onset of voluntary relaxation. SICI decreased in the 80–100 ms before RRT, and MEPs were significantly greater in amplitude in the 60–80 ms period before RRT than in the other intervals in single-pulse trials. TMS pulses did not effectively increase RRT. These results show that cortical excitability in the early stage, before muscle relaxation, plays an important role in muscle relaxation control. SICI circuits may vary between decreased and increased activation to continuously maintain muscle relaxation during or after a relaxation response. With regard to M1 excitability dynamics, we suggest that SICI also dynamically changes throughout the muscle relaxation process.

Exposure of ornithine decarboxylase (ODC;L-ornithine carboxy-lyase, EC 4.1. We isolated ODC-overproducing mouse FM3A cells termed EXOD-1 (17). Since exposure of these ODCoverproducing cells to micromolar levels of spermine or spermidine caused abnormal accumulation and toxicity of polyamines and since overproduction of ODC may deplete antizyme levels, we tried to determine if antizyme is a negative regulator of polyamine transport that inhibits the overaccumulation of polyamines, using EXOD-1 cells transfected with antizyme cDNA. In fact, it was clearly demonstrated that antizyme negatively regulates polyamine uptake. MATERIALS AND METHODSCell Culture and Transfection. ODC-overproducing mouse FM3A cells (EXOD-1) were previously isolated as described (17). The cells (2 x 104 cells per ml) were cultured in ES medium (Nissui Pharmaceutical, Tokyo) supplemented with 50 units of streptomycin and 100 units of penicillin G per ml and 2% (vol/vol) heat-inactivated fetal calf serum (FCS) at 370C in an atmosphere containing 5% CO2. The Mg2+ concentration in the medium was adjusted to 0.9 mM, since Mg2+ has been found to influence polyamine transport (18).pMAMneoZl, possessing the majority (93%) of the antizyme coding region downstream from the promoter of the dexamethasone-inducible mouse mammary tumor virus long terminal repeat, was prepared as described (12). Transfection of EXOD-1 cells with pMAMneoZl was performed by the calcium phosphate coprecipitation method according to the manufacturer's instructions (Stratagene) with DNA at 20 pg/ml per 2.5 x 106 cells on a 10-cm plate, and cells were cultured overnight in ES medium containing 0.5% FCS and the above antibiotics. Cells were cultured further in ES medium containing 2% FCS and the antibiotics for 24 h. Cell lines were cloned from foci isolated after additional growth for 10 days in 0.5% soft agar containing ES medium, the antibiotics, 5% FCS, 5 mM a-difluoromethylornithine (DFMO), and 1 mg ofG418 (geneticin) per ml, and -80 clones containing pMAMneoZ1 were isolated. ODC-overproducing EXOD-1 cells and a pMAMneoZl transfectant (AZ-2) were cultured in ES medium containing 2% FCS and 5 mM DFMO for 24 h in the presence and absence of 1 ,uM dexamethasone, and the effects of spermine and spermidine on cell growth were then examined in the presence of 1 mM aminoguanidine, an inhibitor of amine oxidase in serum (19). DFMO was kindly provided by Marion Merrell Dow (Cincinnati).Assay for Polyamine Transport. After washing FM3A cells with NaCl buffer (135 mM NaCl/1 mM MgCl2/2 mM CaCl2/10 mM glucose/20 mM Hepes, pH adjusted to 7.2 with Tris), the polyamine transport activity was measured as described (6)

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