Combined pre- and postsynaptic action of IgG antibodies in Miller Fisher syndrome B. Buchwald, MD*; J. Bufler, MD*; M. Carpo, MD, PhD; F. Heidenreich, MD; R. Pitz, PhD; J. Dudel, MD; E. Nobile–Orazio, MD, PhD; and K.V. Toyka, MD Article abstract—Background: Miller Fisher syndrome (MFS), a variant of the Guillain-Barré syndrome, is associated with the presence of neuromuscular blocking antibodies, some of which may be directed at the ganglioside GQ1b. Materials and Methods: The authors investigated the in vitro effects of serum and purified immunoglobulin (Ig) G in a total of 11 patients with typical MFS during active disease, and in three of those patients after recovery. From one patient’s serum, we prepared an IgG fraction enriched in anti-GQ1b antibodies by affinity chromatography. For combined pre- and postsynaptic analysis, endplate currents were recorded by a perfused macro-patch clamp electrode. Postsynaptic nicotinic acetylcholine receptor channels were investigated by an outside-out patch clamp technique in cultured mouse myotubes. Results: All MFS-sera depressed evoked quantal release and reduced the amplitude of postsynaptic currents. Five of the 11 sera were additionally examined by outside-out patch clamp analysis and caused a concentration-dependent and reversible decrease in acetylcholine-induced currents. The time course of activation and desensitization of nicotinic acetylcholine receptor channels was not altered by MFS-IgG. Nine patients (82 %) were positive for anti-GQ1b antibodies in ELISA and dot– blot. The enriched anti-GQ1b antibody fraction had a similar effect as whole serum. After recovery from MFS, blocking activity was lost and sera originally positive for anti-GQ1b antibodies became negative. Conclusion: Circulating IgG antibodies induce both pre- and postsynaptic blockade and may play a pathogenic role in acute MFS. NEUROLOGY 2001;56:67–74 Miller Fisher syndrome (MFS) is characterized by the triad of gait ataxia, external ophthalmoplegia, and areflexia. 1 As a variant of the Guillain–Barré syndrome (GBS), MFS is thought to be an immuno- pathologic disorder of the peripheral nervous system. Immunoglobulin (Ig) G autoantibodies directed at the ganglioside GQ1b are found in the acute phase sera of more than 90% of patients with MFS. 2-4 Gan- gliosides are present in high concentrations in pe- ripheral nerve axons and myelin, and several studies indicated that different gangliosides (e.g., GM1) are present at nodes of Ranvier and at neuromuscular junctions. 5 The distal motor nerve terminal lacks the blood–nerve barrier, making it accessible for circulat- ing antibodies and a suitable site to investigate the functional effects of serum-antibodies experimentally. There is evidence that MFS sera block neuromus- cular transmission in the mouse hemidiaphragm. 6-9 Using intracellular microelectrode techniques, a marked increase in spontaneous release (i.e., minia- ture endplate potential frequency) after application of MFS sera, followed by inexcitability of the muscle upon phrenic nerve stimulation has been demon- strated. 6,9 In that study, the effects depended en- tirely on the presence of complement, 9 and miniature endplate potential (MEPP) amplitudes appeared un- changed. 6,9 With a perfused macro-patch clamp elec- trode, we have previously shown that the purified IgG fraction and fractions containing monovalent, antigen-binding (Fab) fragments of two patients with MFS rapidly and reversibly depressed evoked quan- tal release, whereas the IgM fractions had no ef- fect. 7,8 Furthermore, MFS-IgG reduced the amplitude of single quanta, indicating a superimposed postsyn- aptic effect. 8 In the current study, we addressed the postsynap- tic side more directly by examining the effect of MFS-IgG on acetylcholine (Ach)-induced currents in mouse myotubes using outside-out patch clamp anal- ysis. Using the perfused macro-patch clamp elec- trode we investigated whether the presence of blocking antibodies is a more general feature in pa- tients with MFS, and whether it is disease-related. Materials and methods. Patient sera and purified IgG. Eleven patients with MFS were defined according to standard clinical and electrophysiologic criteria. 10,11 Six men and five women between 23 and 71 years of age were included. One patient had a preceding gastrointestinal infection and six patients had an upper respiratory tract *These authors contributed equally to this work. From the Neurologische Klinik der Bayerischen Julius–Maximilians–Universität Würzburg, Germany; Neurologische Klinik der Technischen Universität München (Drs. Bufler and Pitz), Germany; “Giorgio Spagnol” Laboratory of Clinical Neuroimmunology (Drs. Carpo and Nobile–Orazio), Institute of Clinical Neurology, Centro Dino Ferrari, University of Milan, Italy; Neurologische Klinik der Medizinischen Hochschule Hannover (Dr. Heidenreich), Germany; and Physiologisches Institut der Technischen Universität München (Dr. Dudel), Germany. Supported in part by grants from the Deutsche Forschungsgemeinschaft (Bu-398/3-2), by Gemeinnützige Hertie Stiftung, and by University funds (Klinische Forschungsgruppe für Multiple Sklerose und Neuroimmunologie); also by Associazione Italiana Sclerosi Multipla and Telethon Italy (grant no. 674). Received February 21, 2000. Accepted in final form September 12, 2000. Address correspondence and reprint requests to Dr. Brigitte Buchwald, Neurologische Klinik der Universität Würzburg, Josef-Schneider-Strasse 11, D-97080 Würzburg, Germany; e-mail: brigitte.buchwald@mail.uni-wuerzburg.de Copyright © 2001 by AAN Enterprises, Inc. 67