Fig. 3

Gonadotropin-releasing hormone (GnRH) induces a synchronized increase of calcium in all luteinizing hormone (LH) cells and partial increase in follicle-stimulating hormone (FSH) cells. (A) Top: Image of a dissected head with pituitary exposed from the ventral side of the fish used for the ex vivo assays (OB, olfactory bulb; OC, optic chiasm; PIT, pituitary; TH, thalamus; MO, medulla oblongata). Bottom: A diagram describing the ex vivo setup with a constant flow of artificial cerebrospinal fluid (ACSF), a side tube to inject stimuli, and a collecting tube. (B) A representative analysis of basal calcium activity of LH and FSH cells. The left panel is a heatmap of calcium traces (?F/F), where each line represents a cell, with the mean calcium trace on top, the separated line at the bottom of each heatmap is the calcium trace of the chosen region of interest (ROI). The color-coded data points on the right are superimposed on the imaged cells and represent the maximum cross-correlation coefficient between a calcium trace of an active chosen ROI to those of the rest of the cells in the same population, the matrix on the right represent the maximum cross-correlation coefficient values between all the cells (see Figure 3?figure supplement 2 for additional cell activity parameters, scale bar = 20 µm). (C) An analysis of calcium response to GnRH stimulation in two representative imaging sessions, fish 1 where both LH and FSH cells respond, and fish 2 where only LH respond. (see Figure 4?figure supplement 1 for detailed coefficient values distribution in each fish, scale bar = 20 µm). (D) The mean of max cross-correlation coefficient values in each cell type under each treatment (n=10, 3 males, 7 females, one-way ANOVA, ****p<0.0001, see Figure 4?figure supplement 1A for cell-specific vlaues). (E) The percentage of cells responsive to GnRH stimulus (i.e. coefficient values higher than the 80 percentiles of basal values). Each dot represents one fish (n=10, 3 males, 7 females, unpaired t-test, ***p<0.001).