Pharmacological stimulation of galanin receptor 1 but not galanin receptor 2 attenuates kainic acid-induced neuronal cell death in the rat hippocampus

We now celebrate that it is 30 years since galanin was first isolated. During these three decades galanin has been identified in numerous tissues and physiological processes, and in an abundant number of species. In the nervous system galanin primarily displays a modulatory role. The galaninergic system consists of a number of bioactive peptides with a highlyplastic expression pattern and three different receptors, GalR1-GalR3. The lack of receptor subtype selective ligands and antibodies have severely hampered the characterization of this system. Therefore, most of the knowledgehas been drawn from experiments with transgenic animals, which has givensome major conclusions, despite the risk of inducing compensatory effects inthese animal studies. Therefore, the production of subtype selective ligandsis of great importance to delineate the galanin system and slowly experimental data from receptor subtype selective ligand trials is emerging. This thesis aims at studying galanin receptor-ligand interactions and to increase and improve the utilized tools in the galanin research field, especially the development of novel galanin receptor subtype selective ligands. Paper I demonstrates the potential to N-terminally extend galanin analogues and the successful development of a GalR2 selective ligand. In addition, a cell line stably expressing GalR3 was developed to improve and simplify future evaluations of receptor subtype selective galanin ligands. Paper II extends the number of GalR2 selective ligands and shows that i.c.v. administration of galanin receptor ligands stimulates food intake through GalR1. Paper III demonstrates the successful development of a mixed GalR1/GalR2 agonist without any detectable interaction with GalR3. Subsequently, this peptide was used to delineate which receptor subtype mediatesthe neuroprotective effects of galanin in the CA3 region of hippocampus. Furthermore, a robust protocol for detection of receptor activation was developed to ease the detection of the relative potency of novel ligands at the three galanin receptor subtypes. Paper IV describes the finding of several essential amino acids for ligand interaction in GalR3 through the performance of an L-alanine mutagenesis study. A constructed in silico homology model of GalR3 confirmed and extended these findings. In conclusion, this thesis provides a novel design strategy for galanin receptor ligands and increases the understanding of ligand interactions with the GalR3. Furthermore, published ligands together with new galanin analogues have proven to be highly receptor specific, thus implicating that a future delineation of the galaninergic system as a therapeutic target is possible.