Differential intracellular staining of identified neurones in Locusta with texas red and lucifer yellow.

The bright red fluorescent dye, Texas red, is introduced for ionophoretic microinjection in conjunction with the well-known dye Lucifer yellow. Different identified neurones can thus be visualised in two strongly contrasting colours in the same preparation (differential intracellular staining) following their physiological characterisation. Satisfactory results were obtained with electrodes filled with 4% Texas red (sulforhodamine 101 acid chloride; w/v) in 1 M potassium acetate (pH 3.0) and 5% Lucifer yellow (w/v) in aqua dest., respectively. Texas red was injected ionophoretically with pulsed depolarising current (3-10 nA, 500 ms pulses at 1 Hz, 15-30 min) and Lucifer yellow with hyperpolarizing constant current (5-6 nA, 5-15 min). Histological tissue processing was identical for both dyes, the quality of intracellular recordings with Texas red electrodes was similar to that with Lucifer yellow electrodes. Stained neurones could be visualised in both whole-mounts and sectioned preparations. Differential staining of two identified synaptically coupled neurones, a motoneurone and an interneurone, in the mesothorax of Locusta is presented as an illustration for the possible localisation of contact sites at the light-microscopic level.