Recent study showed that cannabinoid CB1 receptor-mediated signaling may take a huge role in age-related modification of brain function. CB1 deficient animals show significant cognitive deficit in old age, however they show better learning ability in young age. Even though the finding is surprising, the molecular mechanisms of such difference, especially for better learning in young age remains unknown mainly due to lack of suitable in-vivo/ex-vivo models. Here we will purpose to use the primary cultured neurons as the ex-vivo model of aging, which can be enabled with gentleMACS technology. This method is said to enable the primary cell culture from adult rodents but up to age of P60. In the present study we aimed to establish a method for primary culture of brain neurons using aged mice, which can be also used for knockout animals of CB1 receptors.
We first tried to prepare the primary culture of brain neurons from neonatal (P7), young (P50) and mature (P105) mice. After 3-days of culture we could find that all of them survived well and start to extend their axons. Notably, P105 is twice an older age than previously reported suggesting that this method could be usable to evaluate the neuronal growth and cellular activity after the certain aging. Further, they could even survive for next 10-days with continuous axon growth. Using this method, we will further conduct the primary cell culture in knockout animals of CB1 as well as endocannabinoid producing enzyme DAGL-a to see the effect of endocannabinoid deficiency to the neuronal growth and cellular activity.