Terpenes control Chlorodiazepoxide-membrane interaction by membrane dipole potential

TURINA A. DEL V.; PERILLO A.M.

Instituto Butantan – San Pablo, Brasil

Workshop; International Workshop on Spectroscopy for Biology.; 2001

  In previous works we demonstrated neurotrophic effects of the essential oil from Tagetes minuta and a relationship with specific and nonspecific benzodiazepine-synaptosomal membrane interactions from brain cortex and with the modulation of the supramolecular coupling of GABAA receptor (GABAA-R) subuinits through a mechanism involving terpenes incorporation into membranes. In the present work we investigated the ability of several natural terpenes to affect the localization of the benzodiazepine chlorodiazepoxide (CDX) within biomembranes, using micelles of a neutral detergent as a membrane model with or without cineol, thymol, menthol, geraniol and comphor. Chlorodiazepoxide was chosen due to its relatively high pK value. All the terpenes assayed induced an increment on the value of CDX’ pK respect to the one obtained in the presence of pure Triton X-100 micelles (pKapp). Negative dipole potentials (j between -171 to –306 mV repect to pure detergent) were obtained when a Boltzman equation was applied to correct Kapp, as an attempt to look for a justification of the difference between pKi in the presence of Triton X-100 with or without terpenes which was too big to be ascribed to a mere polarity change in the microenvironment. This suggested an increased tendency of the species CDXH+ to remain in the membrane phase.  Merocyanine was also used as an electrochromic dye. From the value of lmax of merocyanine monomer, dielectric constant (D) of the microenvironment of this negative dye was obtained in the presence of Triton X-100 (D=9) and decreased in the presence of all the terpenes assayed (D= 10.9-12). As partitioning of merocyanine had been associated to surface potential, a second set of negative j values (betwenn –9 and –20 mV) were calculated from A570. Changes of surface charge density (Ds) upto a 30% due to the partition of the negative charged dye molecules should not be neglected. Taking into account this fact, both j values (calculated from DpK of CDX and from partitioning of merocyanine) may be considered of similar magnitude and of dipolar origin. The latter is due to the fact that neither Triton x-100 nor the terpenes assayed exhibit net charges within the whole pH range tested. Our results were compatible with a model consisting of terpenes molecules with a dipole moment oriented anti-parallel to the intrinsic dipole of the Triton X-100 molecules in the micelles, leading to a more negative environment of the interface region where CDX is located and decresing the net polarity of the deepest micelle regions sensed by merocyanine. The hypothesis that modulatory effects of terpenes on GABAA-R activity could be exerted by subtle changes in the dipolar organization of its environment is discussed.