This experiment was conducted for the development of induced breeding techniques of Neotropius atherinoides using pituitary gland (PG) extract at the Floodplain Substation, BFRI, Santahar, Bogura, Bangladesh. Three breeding trials were conducted during the months of April, May, and June where fifteen pairs of males and females were used for each trial. During the first trial in April, PG extract was used at a rate of 25, 30, and 35mg PG/ kg body weight of the female fish; however, no fish individual was ovulated. Then, two more breeding trials were conducted in May and June using 12 mg (T₁), 13 mg (T₂), and 14 mg (T₃) PG/ kg body weight of females and 6 mg (T₁), 7 mg (T₂), and 8 mg (T₃) PG/ kg body weight of male fish. During this time, the ovulation rates were recorded as 63.12±1.21, 65.21±2.13, and 69.14±1.59% in May and 65.23±1.62, 68.59±1.80, and 71.87±1.91% in June under T₁, T₂ and T₃ treatments, respectively. Fertilization rates of eggs were estimated as 59.24±1.12, 61.36±1.31, and 62.34±1.21% in May and 62.85±1.30, 64.74±1.51, and 66.39±1.40% in June, respectively. Hatching rates of eggs were recorded as 75.54±2.11, 79.64±1.62, and 80.33±1.10% in May and 80.89±1.30, 82.48±1.12, and 86.98±1.90% in June under T₁, T₂ and T₃ treatments, respectively. Significantly (P<0.05) higher values of the mean gonado-somatic index were observed during June, July, and August for females and July showed the highest value. The female and male showed the best breeding performance in June treated with the dose of 14mg and 8mg PG/ kg body weight, respectively.  Results from the present experiment revealed that induced breeding of N. atherinoides, using PG extract is successful which might be helpful in mass seed production of this species for aquaculture as well as conserving the species in nature.