Lambert-Eaton sera reduce low-voltage and high-voltage activated Ca2+ currents in murine dorsal root ganglion neurons.

Abstract

Voltage-gated Ca2+ channels are categorized as either high-voltage activated (HVA) or low-voltage activated (LVA), and a subtype (or subtypes) of HVA Ca2+ channels link the presynaptic depolarization to rapid neurotransmitter release. Reductions in transmitter release are characteristic of the autoimmune disorder, Lambert-Eaton syndrome (LES). Because antibodies from LES patients reduce Ca2+ influx in a variety of cell types and disrupt the intramembrane organization of active zones at neuromuscular synapses, specificity of LES antibodies for the Ca2+ channels that control transmitter release has been suggested as the mechanism for disease. We tested sera from four patients with LES. Serum samples from three of the four patients reduced both the maximal LVA and HVA Ca2+ conductances in murine dorsal root ganglion neurons. Thus, even though LES is expressed as a neuromuscular and autonomic disorder, our studies suggest that Ca2+ channels may be broadly affected in LES patients. To account for the specificity of disease expression, we suggest that incapacitation of only a fraction of the Ca2+ channels clustered at active zones would severely depress transmitter release. In particular, if several Ca2+ channels in a cluster are normally required to open simultaneously before transmitter release becomes likely, the loss of a few active zone Ca2+ channels would exponentially reduce the probability of transmitter release. This model may explain why LES is expressed as a neuromuscular disorder and can account for a clinical hallmark of LES, facilitation of neuromuscular transmission produced by vigorous voluntary effort.Lambert-Eaton myasthenic syndrome (LES) is a human autoimmune disorder characterized by autonomic dysfunction and muscle weakness resulting from reduced neurotransmitter release (1, 2). At neuromuscular junctions, transmitter release occurs at discrete sites called active zones (3, 4). In freezefracture views of neuromuscular synapses, intramembrane particles, which are thought to include Ca2+ channels, are organized in linear arrays at active zones (5-7). The hypothesis that LES antibodies target the Ca2+ channels that control transmitter release arose from the observation that LES antibodies decrease the number of active zones and disrupt active zone organization in mice and humans (8-10). Further support for this hypothesis has been based on the demonstration that LES antibodies decrease Ca2+ influx in transformed cells such as small cell lung carcinoma (11, 12), rat (13) and human (14) MATERIALS AND METHODS Serum Preparation. Serum obtained from four LES patients was dialyzed (exclusion of .100 kDa) for 24 hr against culture medium at a sample-to-dialysate ratio of approximately 1:100, with one dialysate change at about 8 hr (12). Initial experiments for Patient One used whole serum without dialysis; because no obvious differences were observed between whole and dialyzed serum, the results for this patient were pooled and averaged. Serum was added to the culture medium at approximately 1:20 ...

Reduction of Calcium Currents by Lambert–Eaton Syndrome Sera: Motoneurons Are Preferentially Affected, and L-Type Currents Are Spared

“…The cells were plated on 35 mm dishes that had been coated overnight by exposure to poly-L-lysine (4 -15 kDa; 1 mg/ml in 0.15 M boric acid, pH 8.4). Serum from either normal humans or one of the LES patients was dialyzed (exclusion of Ն100 kDa) for 24 hr against culture medium at a sample/ dialysate ratio of ϳ1:100 with one dialysate change at ϳ8 hr (García et al, 1996). Dialyzed serum was added to the culture medium at ϳ1:20 dilution at the time of cell plating.…”

“…Comparatively, these sera cause a lesser decrease in calcium currents in DRG neurons (previously reported by García et al, 1996) and have little effect on muscle. Within motoneurons, voltage-gated potassium channels are not affected, both LVA and HVA calcium currents are decreased, and the HVA calcium current remaining in LES serum-treated motoneurons is mostly L-type.…”

“…In addition to comparing current densities at specific test potentials, we also fitted the peak current-voltage relationship of individual cells as the sum of current through two populations of channels, each activating in accord with a Boltzmann function (García et al, 1996). These fits yielded a maximal conductance of the LVA and HVA components of current, which were averaged for each group of cells (Fig.…”

“…To test this hypothesis, the effects of LES antibodies on radiocalcium fluxes or calcium currents have been examined in a number of systems. The antibodies reduce calcium influx in small-cell lung carcinoma (Roberts et al, 1985), rat anterior pituitary (Login et al, 1987), adrenal chromaffin (Kim and Neher, 1988;Viglione et al, 1992), neuroblastoma (Peers et al, 1990;Grassi et al, 1994), rat thyroid cell line and dorsal root ganglion (DRG) cells (García et al, 1996). Recently, it has been shown that LES antibodies decrease mixed sodium and calcium currents at the nerve terminals of mice (Smith et al, 1995).…”

Anti-Ca 2+ channel antibody attenuates Ca 2+ currents and mimics cerebellar ataxia in vivo

“…Our work establishes the central pathogenicity of antibody against the D-III region. The high homology between P/Q-and N-type VGCCs in this region supports one explanation for the ability of sera from LEMS patients to inhibit more than one type of high-voltage-activated Ca 2ϩ channels: the antibodies target a well conserved epitope (27). In contrast, antibody against D-IV (22, 23), another epitope commonly found in LEMS patients (28), neither reduced Ca 2ϩ influx into parallel fiber terminals in cerebellar slices nor caused loss of motor ability in vivo.…”