Rodrigues, Aldo Rogelis A. and Donata Oertel. Hyperpolarizationactivated currents regulate excitability in stellate cells of the mammalian ventral cochlear nucleus. J Neurophysiol 95: 76 -87, 2006. First published September 28, 2005 doi:10.1152/jn.00624.2005. The differing biophysical properties of neurons the axons of which form the different pathways from the ventral cochlear nucleus (VCN) determine what acoustic information they can convey. T stellate cells, excitatory neurons the axons of which project locally and to the inferior colliculus, and D stellate cells, inhibitory neurons the axons of which project to the ipsi-and contralateral cochlear nuclei, fire tonically when they are depolarized, and, unlike other cell types in the VCN, their firing rates are sensitive to small changes in resting currents. In both types of neurons, the hyperpolarization-activated current (I h ) reversed at -40 mV, was activated at voltages negative to Ϫ60 mV, and half-activated at approximately Ϫ88 mV; maximum hyperpolarization-activated conductances (g h max ) were 19.1 Ϯ 2.3 nS in T and 30.3 Ϯ 2.6 nS in D stellate cells (means Ϯ SE). Activation and deactivation were slower in T than in D stellate cells. In both types of stellate cells, 50 M 4(N-ethyl-N-phenylamino)1,2-dimethyl-6-(methylamino) pyridinium chloride (ZD7288) and 2 mM Cs ϩ blocked a 6-to 10-fold greater conductance than the voltage-dependent g h determined from Boltzmann analyses at -62 mV. The voltageinsensitive, ZD7288-sensitive conductance was proportional to g h max and g input . 8-Br-cAMP shifted the voltage dependence of I h in the depolarizing direction, increased the rate of activation, and slowed its deactivation in both T and D stellate cells. Reduction in temperature did not change the voltage dependence but reduced the maximal g h with a Q 10 of 1.3 and slowed the kinetics with a Q 10 of 3.3.

1 We investigated the action of TsTX-Ka on cloned Kv1.3 channels of the Shaker subfamily of voltage-gated potassium channels, using the voltage -clamp technique. Highly purified TsTX-Ka was obtained from the venom of the Brazilian scorpion Tityus serrulatus using a new purification protocol. Our results show that TsTX-Ka blocks Kv1.3 with high affinity in two expression systems. 2 TsTX-Ka blockade of Kv1.3 channels expressed in Xenopus oocytes was found to be completely reversible and to exhibit a pH dependence. The K D was 3.9 nm at pH 7.5, 9.5 nm at pH 7.0 and 94.5 nm at pH 6.5. 3 The blocking properties of TsTX-Ka in a mammalian cell line (L929), stably transfected to express Kv1.3, were studied using the patch -clamp technique. In this preparation, the toxin had a K D of 19.8 nm at pH 7.4. 4 TsTX-Ka was found to affect neither the voltage-dependence of activation, nor the activation and deactivation time constants. The block appeared to be independent of the transmembrane voltage and the toxin did not interfere with the C-type inactivation process. 5 Taken as a whole, our findings indicate that TsTX-Ka acts as a simple blocker of Kv1.3 channels. It is concluded that this toxin is a useful tool for probing not only the physiological roles of Kv1.2, but also those mediated by Kv1.3 channels.

Aim:The increase in the number of fungal infections worldwide, coupled with the limitations of current antifungal chemotherapy, demand the development of safe and effective new antifungals. Here, we presented the synthesis of a novel acridone (M14) and its antifungal properties against Candida and dermatophytes species. Methods and Results: A series of 17 acridones was designed, synthesized and tested for its antifungal activity. The minimum inhibitory concentration (MIC) was determined by the broth microdilution method. Only the acridone M14 showed growth-inhibitory activity against reference strains and clinical isolates of Candida and dermatophytes, with MIC range of 7Á81-31Á25 µg ml À1 . Moreover, M14 exhibited fungicidal activity and prevented biofilm formation by C. albicans as well as reduced the viability of preformed biofilms, even at sub-MICs. The confocal laser scanning microscopy analysis revealed that C. albicans hyphal growth was completely inhibited in the presence of M14. Similarly, there was a severe inhibition on hyphal growth of Trichophyton rubrum. We also found that M14 has relatively low toxicity to human fibroblasts. Conclusions: The new acridone M14 has antifungal properties against Candida spp. and dermatophytes, and antibiofilm activity against C. albicans. In addition, M14 is relatively selective to fungal cells compared to human normal cells. Significance and Impact of the Study: Because of its in vitro antifungal activity, anti-Candida biofilm effect and moderate cytotoxicity towards normal human cell, M14 may serve as a valuable lead compound to develop a new antifungal agent.

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