C-terminal fragment of tetanus toxin heavy chain modulates sphingomyelinase activity in neuronal preparations

Tetanus toxin (TeTx) reaches CNS by means of retrograde transport inside motor neuron axons. The C-terminal domain of TeTx (HC-TeTx) has been described as responsible for binding and internalization of the toxin into neurons. Analysis of this transport reveals common endocytic carriers for HC-TeTx and neurotrophin signalling (i.e. p75NTR, TrkB, NGF and BDNF). Moreover, in previous work from our group, the activation of neurotrophic signalling by HC-TeTx was described. On the basis of these observations, we decided to study the influence of HC-TeTx in p75NTR-dependent events. Since p75NTR lacks intrinsic enzymatic activity and has been described as a sphingomyelinase (SMase)-coupled receptor, the modulation of SMase activity after HC-TeTx treatment in cultured cerebellar granule neurons (CGN) has been determined and compared with the action of NGF. Neutral- and acid-sphingomyelinases are enzymes that catalyze the hydrolysis of sphingomyelin into ceramide and phosphorylcholine. Treatments of CGN at 7 DIV show a moderate yet consistent and significant enhancement of the nSMase activity after HC-TeTx, as assessed by means of enzymatic assays. HC-TeTx treatment of 14C-serine-labelled CGN and determination of ceramide levels after subsequent thin-layer chromatography show an increase in the ceramide/sphingomyelin ratio. These results shed more light on the common events that are shared by neurotrophins and TeTx.