Methods, materials and procedures for producing viable hypothalamic slices are described in detail. Also described are the results of methodological experiments dealing with combatting the problem of evaporative water loss which produces subsequent increases in concentration of the bathing medium. A formula is given by which the amounts of evaporative loss may be calculated and compensated for without direct measurement of the medium osmotic pressure. Finally, ultrastructural data are presented which indicate that paraventricular nucleus neurosecretory cells in the slices undergo a loss of dense core vesicles during the initial 3 hr in vitro, then recover by 5 hr and maintain a relatively constant state for up to 9 hr, the longest time sampled.