Memory decline is considered a normal part of the aging process, while the relationship between melatonin deficit and cognitive functions is complex and not fully understood. The present study explored the role of melatonin deficiency induced in different age stages on changes in recognition and spatial memory in rats. Age-related worsening of memory learning capacity, working and short-term memory, tested in the Y maze, object recognition test, and radial arm maze was detected. Pinealectomy negatively impacted memory capacity in the young adult and middle-aged rats, while old rats were unaffected. The reduced ratio of pCREB/CREB in the frontal cortex and hippocampus was associated with memory impairment in young adult and middle-aged rats with melatonin deficiency. While aging reduced the expression of Brain-Derived Neurotrophic Factor (BDNF) in the whole hippocampus, pinealectomy exacerbated this process in the CA3 region of 3- and 14-month-old rats with pinealectomy. Changes in the expression of Extracellular Signal-Regulated Kinase (ERK)1/2 and pERK 1/2 were age- and region-specific. The vulnerability of the young adult and middle-aged rats with melatonin deficiency to memory decline suggests a critical period for intervention. Our study provides insights into the molecular pathways involved in age-related memory changes associated with melatonin deficiency, emphasizing the importance of the BDNF/CREB/ERK1/2 pathway in the hippocampus.Acknowledgements: This work was supported by the National Science Fund of Bulgaria (research grant # KΠ -06-H41/1, 30 November 2020).