No abstract

—Herpes simplex virus — type 1 (HSV‐I) plaque‐forming ability and plaque size were measured on C3H/1OT1/2 cell monolayers as functions of pretreatment dose with UV light at different times before inoculation with virus, in order to determine if UV‐enhanced reactivation (ER) of UV‐irradiated virus. as well as associated phenomena, could be obtained in this cell system. The number of virus plaques observed (i.e. the capacity of the cells to support virus growth) and the size of the plaques were found to increase substantially with pretreatment of the cells with UV light. However, no significant ER was observed. Therefore, the mechanisms responsible for the increases in plaque size and cell capacity seem to be independent of those responsible for ER. In work by others. C3H/l0T1/2 cells have hcen transformed by UV light at doses similar to those used in this study; the absence of ER of UV‐irradiated virus in this study indicates that the mechanism underlying ER is not required for transformation.

Stable isotopes of Mg were used to measure bidirectional magnesium ion fluxes in single barnacle giant muscle fibers immersed in Ca- and Na-free, isosmotic media. Measurements were made using a mass spectrometric technique, thermal ionization mass spectrometry (TIMS), in conjunction with atomic absorption spectroscopy. Kinetic relations based on a first-order model were developed that permit the determination of unidirectional rate coefficients for Mg influx, ki, and efflux, ke, in the same experiment from knowledge of initial conditions and the initial and final ratios of 26Mg/24Mg and 25Mg/24Mg in ambient solutions (i.e., by isotope dilution). Such determinations were made for three values of the external Mg ion concentration: 5, 25, and 60 mM. At the concentration [Mg+2]o = 5 mM, ki and ke were about equal at a value of 0.01 min-1. At the higher values of [Mg+2]o, the values of ke increased along a curve suggesting saturation, whereas the values of ki remained essentially constant. As could be expected on the basis of a constant ki, the initial influx rate varied in direct linear proportion to [Mg+2]o, and was 11.8 pmol/cm2s when [Mg+2]o was 5 mM. However, the initial efflux rate appeared to increase nonlinearly with [Mg+2]o, varying from 13.4 pmol/cm2s ([ Mg+2]o = 5 mM) to approximately 80 pmol/cm2s ([ Mg+2]o = 60 mM). The results are consistent with a model that assumes Mg influx to be mainly an electrodiffusive inward leak with PMg = 0.07 cm/s and Mg efflux to be almost entirely by active transport processes. Where comparisons can be made, the rate coefficients determined from stable isotope measurements agree with those previously obtained using radioactive Mg. The rate coefficients can be used to correctly predict time-dependent changes in total fiber Mg content. The results support the conclusion that nonradioactive tracers can be used to measure ion fluxes and ion flux ratios in excitable cells; it is expected that this method will greatly assist in the study of Mg regulation in general.

Rabbit erythrocyte ghosts were fused by means of electric pulses to determine the electrofusion thresholds for these membranes. Two protocols were used to investigate fusion events: contact-first, and pulse-first. Electrical capacitance discharge (CD) pulses were used to induce fusion. Plots of fusion yield vs peak field strength yielded curves that intersected the field strength axis at positive values (pseudothresholds) which depended on the protocol and decay half time of the pulses. It was found that plots of pseudothreshold vs reciprocal half time were linear for each protocol; when extrapolated to reciprocal half time = 0 (i.e., t----infinity), these lines intersected the ordinate at values of the field strength considered to be the true electrofusion thresholds. In this fashion, the contact-first protocol gave an electrofusion threshold of 46.5 +/- 11.5 V/mm for hemoglobin-free ghosts (white ghosts) and 40.9 +/- 8.8 V/mm for ghosts with fractional hemoglobin (pink ghosts), while the threshold for the pulse-first protocol applied to pink ghosts was determined to be 93.4 +/- 11.0 V/mm. Although the thresholds depended on the electrofusion protocol, plots of critical field strength vs reciprocal time had the same slopes, i.e., approximately 24 Vs/mm. The results suggest that the fusogenic state induced by an electric pulse in either the contact-first protocol or the pulse-first protocol (long-lived fusogenic state) may in fact share a common mechanism, if the two states are not actually identical.

Giant axons from the marine annelid Myxicola infundibulum were internally dialyzed with solutions containing 22Na ions as tracers of Na efflux. In experiments performed in Li-substituted seawater, Na efflux that is dependent on external Ca ion concentration, [Ca2+]o, was measured using dialysis to maintain [Na+]i at 100 mM, which enhances the [Ca2+]o-dependent Na efflux component, (i.e., reverse-mode Na/Ca exchange). When dialysis fluid contained EGTA (1 mM) to buffer the internal Ca concentration, [Ca2+]i, to desired levels, Na efflux lost its normal sensitivity to external calcium. The inhibition was not simply due to the Ca-chelating action of EGTA to produce insufficient [Ca2+]i to activate Na/Ca exchange. The addition of EGTA inhibited Cao-dependent Na efflux even when a large enough excess of [Ca2+]i was present to saturate the EGTA and still produce elevated values of [Ca2+]i. Control experiments showed that these high values of [Ca2+]i resulted in normal Na/Ca exchange in the absence of EGTA. It is concluded that the presence of EGTA itself interferes with the manifestation of reverse-mode Na/Ca exchange in Myxicola giant axons.