Dibenzyl trisulphide (DTS), a main lipophilic compound in Petiveria alliacea L. (Phytolaccaceae), was identified as one of the active immunomodulatory compounds in extracts of the plant. To learn more about its biological activities and molecular mechanisms, we conducted one-dimensional NMR interaction studies with bovine serum albumin (BSA) and tested DTS and related compounds in two well-established neuronal cell-and-tissue culture systems. We found that DTS preferentially binds to an aromatic region of BSA which is rich in tyrosyl residues. In SH-SY5Y neuroblastoma cells, DTS attenuates the dephosphorylation of tyrosyl residues of MAP kinase (erk1/erk2). In the same neuroblastoma cell line and in Wistar 38 human lung fibroblasts, DTS causes a reversible disassembly of microtubules, but it did not affect actin dynamics. Probably due to the disruption of the microtubule dynamics, DTS also inhibits neuroblastoma cell proliferation and neurite outgrowth from spinal cord explants. Related dibenzyl compounds with none, one, or two sulphur atoms were found to be significantly less effective. These data confirmed that the natural compound DTS has a diverse spectrum of biological properties, including cytostatic and neurotoxic actions in addition to immunomodulatory activities.

The time course of the effect and the concentration-response relationship were successfully described by a pharmacokinetic/pharmacodynamic model that takes into account the rapid development of tolerance of pupillary light reflex parameters. This provides a basis for further investigations of the applicability of pupillography as a surrogate measurement of the effectivity of antidepressant drugs with norepinephrine reuptake-inhibiting properties.

In SH-SY5Y neuroblastoma cell bodies and growth cones the actin dynamics of cholinergically induced lamellar protrusion is demonstrated by live-imaging. Failure of this reaction in cells over-expressing a dominant negative RAC1-mutant (RAC1(T17N)) confirmed that the actin-dynamics of lamellar protrusion is also, in these cells, RAC1-dependent. Pretreatment of untransfected cells with 20 microM UO126 for 2 h, shown to down-regulate the basic level and to completely inhibit cholinergic activation of mitoge-activated kinase (MAPK; erk1/erk2), had no effect neither on spontaneous nor on induced lamellar protrusions. When p-JNK, whose basic activity is very low and not enhanced by a cholinergic stimulation of the cells for up to 30 min, is activated in the presence of 50 microM anisomycin, both spontaneous and cholinergically induced lamellar protrusive activity is attenuated. These data indicate that the RAC1-controlled cascades promoting lamellar protrusion are independent of MAPK activity and partially down-regulated by p-JNK.

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