The transport rates of three indicators (Ethidium Bromide (EB), Propidium Iodide (PI) and Calcein) across electroporated NIH3T3 cells was studied to gain insight on the resealing kinetics of field induced membrane pores. Additionally, a number of factors that influence the resealing process were examined. In low salt medium, the influx rates (EB and PI) can be described by a single diffusive process while in isotonic high salt medium two distinct processes are identified; namely, a relatively large and rapid influx within a few hundred msec after the pulse followed by a smaller and slow influx at longer times. The observed influx rates in both buffer mediums were dependent on the external dye concentrations. The net influx (total integrated fluorescence) in the presence or absence of either Ca(II) (1.8 mM), or a series of non-ionic synthetic surfactants (P188, F68, T1107) (approximately 1 mg/ml) was assessed using Calcein. A significant decrease in the net influx was observed when these additives were present in the low salt medium, but had little effect when added to the high ionic strength medium. The effect of various inositol phosphates on the frequency of histamine induced Ca(II) oscillations in HeLa cells was investigated. Of the various inositol phosphates examined, only the addition of Ins(1,4,5)P3, Ins(1,3,4,5)P4, and Ins(1,3,4,6)P4 transiently increased ongoing Ca(II) oscillations. Inhibition of Ins(1,4,5)P3 3-Kinase by adriamycin blocked Ca(II) oscillations and only single Ca(II) release was observed upon histamine stimulation. Furthermore, the non-metabolizable analogs of IP3 did not alter the frequency of the ongoing oscillations. These results suggest IP4 may play a role in regulating the frequency of oscillations in HeLa cells. Intracellular generation of hydrogen peroxide induced by growth factors peaks in minutes and rapidly decreases back to the unstimulated level. The difference in protein phosphorylation of stimulated cells (NIH3T3 and A431) in the presence and absence of electroporated catalase was examined. Preliminary data shows EGF fails to induce phosphorylation of intracellular signaling proteins such as EGF-R, PLC-gamma, and PI3K in the presence of electroporated catalase.
