Internal K+ and Na+ concentrations and the volume of hypothalamic synaptosomes.

Hypothalamic synaptosomes, prepared by discontinuous sucrose density gradient centrifugation, were suspended in media of varying osmolarity and ionic composition and the internal concentrations of K+ and Na+ were measured. The intrasynaptosomal volume was determined using [14C]inulin, 35SO4 or [14C]sucrose as extracellular markers. When the synaptosomes were suspended in 0.32 M sucrose the distribution volume of [14C]inulin and 35SO4 were similar. However, when the medium contained 140 mM NaCl and 5 mM KCl the 35SO4 space was equal to the total water space while the distribution volume of [14C]inulin corresponded to 35-51% of the total water space. [14C]sucrose distributed in a larger volume than did [14C]inulin, presumably due to intracellular permeation of sucrose. Using inulin as an extracellular marker the synaptosome volume was found to be inversely proportionate to the tonicity of the medium. [14C]inulin was considered a suitable marker of the extrasynaptosomal space and was used when determining intrasynaptosomal K+ and Na+ concentrations. The internal concentration of K+ was considerably higher than the external indicating that the synaptosomes were able to retain K+ against a concentration gradient. The synaptosomes gained Na+ when transferred to media containing 140 mM NaCl. The internal concentrations of K+ and Na+ were unaffected by glucose and elevated temperature and was only moderately changed during 90 min of incubation. The equilibrium potentials for K+ and Na+ were -68 and 13 mV respectively when the medium consisted of 0.03 M sucrose containing 140 mM NaCl, 5 mM KCl. It is concluded that hypothalamic synaptosomes can maintain a high transmembrane concentration gradient for K+ whereas the membrane is rather easily penetrated by Na+.