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The purpose of this work was to map, on the heavy (H) chain of botulinum neurotoxin A (BoNT/A), the regions that bind to mouse brain synaptosomes (snps). We prepared 60 synthetic overlapping peptides that had uniform size and overlaps and encompassed the entire H chain (residues 499 to 1296) of BoNT/A. The ability of each peptide to inhibit the binding of 125I-labeled BoNT/A to mouse brain snps was studied. The binding of 125I-labeled BoNT/A to mouse brain snps was completely inhibited by free unlabeled BoNT/A, but not by unrelated proteins, indicating that the binding of BoNT/A to mouse brain snps was a specific event. Inhibition studies with the individual peptides showed that, on the H(N) domain, inhibitory activities greater than 10% were exhibited, in decreasing order, by peptides 799-817, 659-677, 729-747, 533-551, 701-719, and 757-775. Lower inhibitory activities (between 5.6% and 8.7%) were exhibited by five other peptides, 463-481, 505-523, 519-537, 603-621 and 645-663. The remaining 18 H(N) peptides had little or no inhibitory activity. In the H(C) domain, peptides 1065-1083, 1163-1181 and 1275-1296 had the highest inhibitory activities (between 25% and 29%), followed (10-12% inhibitory activity) by peptides 1107-1125, 1191-1209 and 1233-1251. Two other peptides, 1079-1097 and 1177-1195, had very low (5.8% and 4.9%) inhibitory activities. The remaining 23 H(C) peptides had no inhibitory activity. Inhibition with mixtures of equimolar quantities of the most active 6 peptides of HN, 5 of H(C) or all 11 of H(N) and H(C) revealed that the peptides contain independent non-competing binding regions. Comparison of the locations of the snp-binding regions on the H-subunit with the regions that bind blocking mouse anti-BoNT/A Abs helped explain the protecting ability of these Abs. In the three-dimensional structure of BoNT/A, the snp-binding regions that completely coincide or significantly overlap with the antigenic regions occupy surface locations and most of them reside in the last half of the H(C) domain. But some of the regions reside in the HN domain and might play a role in the translocation event.

ObjectiveThe purpose of this study is to investigate the usefulness of sympathetic skin response (SSR) and heart rate variability (HRV) for the differential diagnosis of patients with dementia with Lewy bodies (DLB).DesignA diagnostic test study.SettingSingle centre in Japan.ParticipantsWe examined 20 patients with probable Alzheimer's disease (AD) diagnosed with NINCDS-ADRDA criteria and 20 with probable DLB diagnosed with the criteria of the third international DLB workshop.MethodsFor the SSR measurement, surface electrodes were used: the active recording electrode was placed on the palm of the hand and the reference electrode was placed on the dorsum of the same hand. SSR was induced by a median nerve electrical stimulation at an amplitude of 20 mA. For the HRV measurement, the A–A intervals were measured twice for 2 min with an interval of 5 min in a sitting position after a rest of 5 min. From the low-frequency power (LF; 0.02–0.15 Hz) and high-frequency power (HF; 0.15–0.50 Hz), the ratio of LF to HF power (LF/HF) was calculated using the maximal entropy method.ResultsSSR and HRV could detect the abnormality of autonomic function in patients with DLB at sensitivities of 85% and 90%, respectively. On the other hand, SSR and HRV detected an abnormality of autonomic function in patients with AD at sensitivities of 15% and 25% (p<0.05). The combination of the SSR and the HRV (double-positive) indicated abnormal autonomic function was recorded in only 1 of 20 patients (5%) with AD. In contrast, this combination indicated autonomic abnormality in 15 of 20 patients with DLB by our criteria (75%).ConclusionsSSR and HRV can be applied to differentiate DLB from AD.

A specific system for antibody removal from blood circulation in myasthenia gravis (MG) patients was devised by use of the immunoadsorbent bound to an acetylcholine receptor (AChR) peptide that was synthesized corresponding to the sequence of residues 183‐200 of the AChR alpha‐subunit (alpha 183‐200), antibodies which prevent the binding of ACh to AChR. The alpha 183‐200 peptide was confirmed to be immunogenic for induction of an animal model of the disease and for reactivity with MG autoantibodies. We then made use of these results for immunoadsorption therapy through the antigen‐antibody reaction on the molecular level, having given patients relief from myasthenic weakness. The greatest care was taken for the selection of an antigenic region in the molecular structure among various myasthenogenic domains of AChR and for the antigenic conformation of synthetic peptide as the adsorbent to react with antibodies raised against the native protein.