Peptides have been shown to be potent regulators of central nervous system (CNS) activity. As such, peptide-based drugs can serve as a highly effective means to treat diseases of the CNS. Despite this potential, peptides have limited capacity to permeate across the blood-brain barrier (BBB). Nevertheless, advancing strategies in peptide drug design, delivery systems, and transporter targeting are emerging, with such potential to overcome the restrictive nature of the BBB. This chapter addresses peptide drug delivery into the CNS, and in vivo evaluation of peptide permeability across the BBB using the dual-artery in situ brain perfusion model coupled with the technique of capillary depletion.