Secretion of luteinizing hormone is decreased when hens start to incubate their eggs and is increased after nest deprivation or hatching of the eggs. The purpose of this study was to determine whether decreased luteinizing hormone (LH) secretion during incubation in the domestic hen is associated with a decrease in hypothalamic chicken gonadotropin-releasing hormone-I (cGnRH-I) mRNA or peptide. A semiquantitative competitive PCR assay was developed to measure cGnRH-I mRNA. Hypothalamic mRNA was quantified as the amount of GnRH cDNA obtained by reverse transcription of cGnRH-I mRNA. The amount of hypothalamic cGnRH-I mRNA was significantly higher in laying than in incubating hens (38.7 ± 10.3 vs. 7.7 ± 1.6 × 10–17 mol cDNA, p = 0.01, n = 8). The hypothalamic GnRH peptide content was not significantly different between laying and incubating hens in either the preoptic area (286.9 ± 24.01 vs. 269.3 ± 29.3 pg, n = 8) or the basal hypothalamus (1.67 ± 0.19 vs. 1.54 ± 0.21 ng, n = 8). Five days after incubating hens were deprived of their eggs, the resulting increase in LH secretion was associated with a significant increase in hypothalamic content of cGnRH-I mRNA (22.8 ± 2.2 vs. 6.7 ± 1.7 × 10–17 mol cDNAp < 0.001, n = 8). These observations suggest that a decrease in the expression of the cGnRH-I gene is a major factor in maintaining depressed LH secretion in incubating dome