Recent studies indicate a critical role for metabotropic glutamate receptor 5 (mGluR5) in the reinstatement of cocaine seeking. However, the signal transduction pathways through which mGluR5s regulate cocaine seeking have not been identified. Here, we show that intra-accumbens shell administration of an mGluR5 (9.0 μm MPEP), but not mGluR1 (50.0 μm YM 298198), antagonist before a priming injection of cocaine (10 mg/kg) attenuated the reinstatement of drug seeking in rats. Consistent with these results, intra-shell microinjection of the mGluR1/5 agonist DHPG (250 μm) promoted cocaine seeking. Intra-shell administration of a phospholipase C (PLC) inhibitor (40.0 μm U73122) or a protein kinase C (PKC) inhibitor (10.0 μm Ro 31-8220 or 30.0 μm chelerythrine chloride) attenuated cocaine seeking. Pharmacological inhibition of PKC in the shell also blocked intra-shell DHPG-induced reinstatement of cocaine seeking. In addition, cocaine priming-induced reinstatement of drug seeking was associated with increased phosphorylation of PKCγ, but not PKCα or PKCβII, in the shell. Cocaine seeking previously was linked to increased phosphorylation of GluA2 at Ser880, a PKC phosphorylation site, which promotes the endocytosis of GluA2-containing AMPA receptors via interactions with Protein Associated with C Kinase (PICK1). The present results indicated that inhibition of PICK1 (100 μm FSC-231) in the shell attenuated cocaine seeking. There were no effects of any drug treatment in the shell on sucrose seeking. Together, these findings indicate that accumbens shell mGluR5 activation promotes cocaine seeking, in part, through activation of PLC and PKCγ. Moreover, the endocytosis of shell GluA2-containing AMPARs during cocaine seeking may depend on interactions with PKCγ and PICK1.