Calcium Channel Agonists Protect against Neuromuscular Dysfunction in a Genetic Model of TDP-43 Mutation in ALS

Abstract

TAR DNA binding protein (TDP-43, encoded by the TARDBP gene) has recently been shown to be associated with amyotrophic lateral sclerosis (ALS), but the early pathophysiological deficits causing impairment in motor function are unknown. Here we expressed the wild-type human gene (wtTARDBP) or the ALS mutation G348C (mutTARDBP) in zebrafish larvae and characterized their motor (swimming) activity and the structure and function of their neuromuscular junctions (NMJs). Of these groups only mutTARDBP larvae showed impaired swimming and increased motoneuron vulnerability with reduced synaptic fidelity, reduced quantal transmission, and more orphaned presynaptic and postsynaptic structures at the NMJ. Remarkably, all behavioral and cellular features were stabilized by chronic treatment with either of the L-type calcium channel agonists FPL 64176 or Bay K 8644. These results indicate that expression of mut-TARDBP results in defective NMJs and that calcium channel agonists could be novel therapeutics for ALS.

Neuroleptics as therapeutic compounds stabilizing neuromuscular transmission in amyotrophic lateral sclerosis

“…We observed an antagonism of T-type Ca 2+ channels in TDP worms and fish, as reported previously for human channels, and found that pimozide preserved NMJ transmission in zebrafish and mouse models of ALS. Similarly, we previously showed that other calcium channel drugs (FPL 64176 and Bay K 8644) suppressed the NMJ phenotype in mutant TDP-43 ALS zebrafish (14). Through facilitation 2+ channels (LVA) were isolated by patch-clamp recording in vivo in mTDP-43 zebrafish motoneurons using a voltage-clamp protocol (depolarizing from -80 mV to +60 mV) (left panel).…”

“…Through paired motoneuron-muscle cell recordings, we assessed the fidelity of synaptic NMJ transmission and the strength of muscle endplate currents, as in our previous works (14,28), in mTDP-43 fish treated or not overnight with pimozide. Compared with wtTDP43, the fidelity of NMJ synaptic transmission was greatly reduced in larvae expressing mTDP-43 ( Figure 5, B-E, P < 0.05).…”

“…Paired motoneuron/muscle recordings were performed following procedures previously described (14). Briefly, paired motor motoneuron/muscle recordings were obtained by perfusing collagenase (1 mg/ml) over the preparation for 10 minutes.…”

“…In mutant SOD1-expressing mice (10), an early retraction or "die-back" of presynaptic motor endings was observed long before the death of motoneurons (11), and this has also been observed in TARDBP mice (12) and in tissue from patients with ALS (13). Expression of either mutant human TARDBP (14) or FUS mRNA in zebrafish (15) resulted in impaired transmission reduced frequency of miniature endplate currents (mEPCs) and reduced quantal transmission at the NMJ. Expression in muscle of a variety of trophic factors in SOD1 mice has been tested for their ability to stabilize the NMJ (16), but these treatments have shown only limited success in mice and humans (17,18).…”

Centipede venoms as a source of drug leads

“…For example, the roscovitine derivative GV-58, a use-dependent Ca V 2.1 agonist, is being developed for treatment of Lambert-Eaton myasthenic syndrome in which progressive muscle weakness accrues due to autoantibody-mediated removal of presynaptic calcium channels at neuromuscular junctions [69,74]. Interestingly, the Ca V 1 agonists Bay K8644 and FPL 64176 both protect against neuromuscular dysfunction in a zebrafish model of amyotrophic lateral sclerosis, whereas roscovitine is ineffective, consistent with a greater role for Ca V 1 channels in stimulating neurotransmitter release at neuromuscular junctions in lower vertebrates [75]. Bay K8644 also protects against neuronal injury in a mouse model of ischemic stroke when delivered 12-24 h after reperfusion [76].…”