Circadian Locomotor Activity Recording
Objective: To assess circadian rhythm restoration following neural transplant by recording free-running locomotor activity rhythms in constant darkness conditions
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Equipment2
Materials1
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Protocol Steps
Create arrhythmic condition via SCN lesion
Lesion the hamster hypothalamic suprachiasmatic nucleus (SCN) to permanently disrupt overt circadian rhythms
Note: This creates the baseline arrhythmic condition in adult hamsters prior to transplantation
View evidence from paper
“overt circadian rhythms are permanently disrupted following lesions of the hamster hypothalamic suprachiasmatic nucleus (SCN)”
Implant fetal SCN graft
Implant brain grafts containing fetal SCN tissue into the lesioned adult hamsters
Note: Grafts are intended to reestablish circadian function in previously arrhythmic animals
View evidence from paper
“implantations of brain grafts containing the fetal SCN reestablish circadian rhythms of locomotor activity in adult hamsters”
Record locomotor activity in constant darkness
Place animals in constant darkness conditions and record free-running locomotor activity rhythms to assess circadian rhythm restoration
Note: Recording continues until circadian rhythm patterns can be established and analyzed
View evidence from paper
“The restoration of free-running rhythms in conditions of constant darkness is correlated with the presence in the graft”
Assess neuropeptide presence in graft
Perform immunocytochemical characterization to identify presence of neuropeptides in the graft including vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), and vasopressin (VP)
Note: Neuropeptide presence is correlated with successful circadian rhythm restoration
View evidence from paper
“The restoration of free-running rhythms in conditions of constant darkness is correlated with the presence in the graft of neuropeptides normally present in the SCN”
Evaluate graft integration with host brain
Examine whether grafts receive retinal input and send efferents into the host brain to assess functional integration
Note: In several recipients, grafts were found to receive retinal input and appeared to send efferents into the host brain
View evidence from paper
“In several recipients, grafts were found to receive retinal input, and appeared to send efferents into the host brain”
Test light entrainment response
Assess whether animals with restored locomotor rhythms can synchronize (entrain) to light intensities
Note: Animals with restored locomotor rhythms failed to synchronize to light intensities to which SCN-intact animals responded
View evidence from paper
“animals with restored locomotor rhythms did not show gonadal regression in the absence of light, and failed to synchronize (entrain) to light intensities”