Ciliate conjugation usually occurs when cells are moderately starved after rapid proliferation. In Blepharisma japonicum, at the stationary phase, a mating pheromone (gamone 1) is secreted by mating type I cells and stimulates mating type II cells to secrete another mating pheromone (gamone 2), which stimulates mating type I cells. Both stimulated cells are transformed to be able to form pairs. We showed previously that the gamone 1 transcription occurs only in the stationary-phase cells, not in logarithmic-phase cells. However, it is still unknown how rapidly cells response to starvation and express gamone 1. Here we show that gamone 1 expression is initiated by sudden artificial starvation, and we determined the time required for the initiation of transcription and secretion of gamone 1 in B. japonicum. Proliferating cells rapidly responded to artificial starvation, approx. 1.5 h at most was necessary for the initiation of the transcription of gamone 1 gene, and approx. 3 h was needed for the secretion of gamone 1. Gamone 1 expression tested in single cells suggests that gamone 1 expression may be correlated with the cell-cycle, most probably with the certain stage of G1 phase in B. japonicum. Our present findings provide an experimental system to investigate the environmental factors involved in the initiation of conjugation.