Melanocortins and the melanocortin-4 receptor (MC4-R) are enriched in the nucleus accumbens, a brain region that has been implicated in the rewarding action of cocaine and other drugs of abuse. In the present study we use a number of rat behavioral models to show that infusion of a melanocortin peptide antagonist into the nucleus accumbens blocks the reinforcing, incentive motivational, and locomotor sensitizing effects of cocaine. We also show that locomotor responses to repeated cocaine exposure are completely blocked in MC4-R null mutant mice and reduced in Agouti mice that overexpress an endogenous inhibitor of melanocortins in the brain. The results also demonstrate that cocaine administration increases the expression of MC4-R in the nucleus accumbens and striatum, and that MC4-R is co-localized with prodynorphin in medium spiny neurons in the nucleus accumbens. Together, these findings indicate that the behavioral actions of cocaine are dependent on activation of MC4-R, and suggest that upregulation of this receptor by drug exposure may contribute to sensitization of these behavioral responses. Modulation of cocaine reward is a novel action of the melanocortin-MC4-R system and could be targeted for the development of new medications for cocaine addiction.

Patients with one form of cicatricial pemphigoid have IgG antibasement membrane autoantibodies against laminin 5 (alpha3beta3gamma2). Although passive transfer of rabbit anti-laminin 5 IgG to neonatal mice has been shown to induce subepidermal blisters that mimic those in patients, it has not been possible to directly assess the pathogenic activity of human autoantibodies in this animal model because the latter do not bind murine skin. To address this question, a disease model in adult mice as well as SCID mice bearing human skin grafts was developed. Adult BALB/C mice challenged with rabbit anti-laminin 5 IgG developed, in a concentration-related fashion, erythema, erosions, and crusts surrounding injection sites, histologic evidence of noninflammatory, subepidermal blisters, and deposits of rabbit IgG and murine C3 in epidermal basement membranes. Anti-laminin 5 IgG also induced subepidermal blisters in: adult complement-, mast cell-, and immuno-deficient mice; adult BALB/C mice pretreated with dexamethasone; and human skin grafts on SCID mice. Alterations did not develop in matching controls challenged with identical amounts of purified normal rabbit IgG or bovine serum albumin. Using this adult mouse model, human skin grafts on SCID mice were challenged with purified IgG from patients with alpha subunit-specific, anti-laminin 5 autoantibodies, or normal controls. Patient (but not control) IgG induced epidermal fragility as well as noninflammatory, subepidermal blisters in grafted human (but not adjacent murine) skin. Moreover, whereas all mice that received patient autoantibodies had anti-laminin 5 IgG in their circulation, deposits of human IgG were present only in the epidermal basement membranes of grafts. Interestingly, these in situ and circulating autoantibodies were predominately of the IgG4 subclass. These studies demonstrate that human anti-laminin 5 autoantibodies are pathogenic in vivo and describe an animal model that can be used to define disease pathomechanisms and biologically important domains within this autoantigen.

Sera from 20 patients with antiepiligrin cicatricial pemphigoid were studied to define the specific reactivity of their IgG autoantibodies. IgG from all patients bound exclusively to the dermal side of 1 mol/L NaCl split skin and immunoprecipitated laminin 5 (alpha3beta3gamma2) from extracts of human keratinocytes (HKs). Immunoblot studies on purified laminin 5 subunits demonstrated that patient IgG bound alpha3 alone in 16 patients. In two patients, IgG autoantibodies were directed predominantly to the gamma2 subunit, yet showed trace reactivity to alpha3 as well. Sera from two patients did not immunoblot any laminin 5 subunits, their IgG presumably immunoprecipitating laminin 5 via a conformational epitope. Sera from patients with alpha3 subunit-specific IgG immunoprecipitated all subunits of laminin 5 as well as polypeptides of 190 and 200 kDa from the conditioned media of HKs. Preclearance studies and experiments utilizing affinity-purified patient IgG demonstrated that the latter signified laminin 6 (alpha3beta1gamma1) that was bound by cross-reactive alpha3 subunit-specific patient IgG. Sera from patients with gamma2 subunit-specific IgG showed no reactivity to laminin 6, except for faint reactivity provided by low levels of their alpha3 subunit-specific IgG. Taken together, these findings indicate that antiepiligrin cicatricial pemphigoid signifies an autoimmune response to subunits present in laminin 5.