VIP neuroprotective effects are mediated by VPAC2 receptors coupled to different transduction pathways

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Prepro-vasoactive intestinal peptide (VIP) mRNA codes for two neuropeptides: VIP and peptide histidine isoleucine (PHI) in rodents or VIP and peptide histidine methionine (PHM) in humans. Two VIP receptors, shared with a similar affinity by pituitary adenylate cyclase-activating polypeptide (PACAP), have been cloned: VPAC₁ and VPAC₂ [28]. PHI binds to these receptors with a lower affinity. Furthermore, PACAP-27 and PACAP-38, but not VIP, bind with high affinity to a specific PACAP receptor called the PAC₁ receptor. VPAC receptors are preferentially coupled to Gas protein that stimulates adenylate cyclase activity and induces cAMP increase [28]. VPAC receptors can also be coupled to Gaq and Gai proteins that stimulate the inositol phosphate / calcium / protein kinase C (PKC) pathways.

In the P0 hamster, the modulating effects of VIP on excitotoxin-induced heterotopias were mimicked by forskolin, PACAP-38 and by a specific VPAC₂ receptor agonist but not by a VPAC₁ agonist, and were blocked by a protein kinase A (PKA) inhibitor. Taken together, these data suggest that VIP and PACAP can attenuate ibotenate-induced heterotopias in newborn hamster and that this effect is mediated by the VPAC₂ receptor utilizing the cAMP-PKA pathway.

In contrast, in the P5 mouse, forskolin had no detectable effect on ibotenate-induced white matter lesions, suggesting that cAMP production was not involved in VIP-induced neuroprotection [26]. Further supporting this hypothesis, stearyl norleucine VIP, a specific VIP agonist that does not activate adenylate cyclase, mimicked VIP neuroprotective effects [26]. A large range of concentrations of PKA inhibitor, calmodulin-dependent PK inhibitor and phosphatidylinositol 3-OH kinase inhibitor had no significant effect on VIP neuroprotection against white matter excitotoxic cystic lesion [29]. In contrast, a PKC inhibitor and a MAPK kinase (or Mek-1) inhibitor abolished VIP protective effects in a dose-dependent manner [29]. In vitro and in vivo studies revealed that VIP elicited in white matter astrocytes PKC activation of PKC but not of MAPK. In addition to a PKC-like activation in white matter cells at the site of injection, VIP also elicited a PKC-like and MAPK-like activation in cortical plate neurons at distance from the site of injection. In neuronal cultures, while VIP and conditioned medium from control astrocytes had no detectable effect on the activation of PKC and MAPK, medium conditioned by astrocytes cultured with VIP induced a significant PKC and MAPK activation [29].

VIP effects on white matter were mimicked with a similar potency by VPAC₂ agonists and PHI but not by VPAC₁ agonists [30, 31]. Surprisingly, VIP-induced neuroprotection was not mimicked by a large range of doses of PACAP 27 or PACAP 38 [27, 31].  This atypical pharmacology of VIP-induced neuroprotection in P5 mice raised several hypotheses: i) activation of PAC₁ receptors could have a toxic effect on the excitotoxic lesions while activation of VPAC receptors could be neuroprotective, leading to a lack of detectable effect for PACAP. In this context, it has been shown that VIP can provide cellular protection through a specific splice variant of the PAC₁ receptor [32]. ii) During some stages of brain development, the binding of VIP or PACAP to VPAC receptors leads to activation of separate transduction pathways. iii) VIP acts through a yet to be identified specific VIP receptor which is not recognized by PACAP. Indeed, Ekblad et al. [33] characterized a PACAP 27 preferring receptor and a VIP specific receptor, distinct from those that have been cloned (VPAC₁, VPAC₂, and PAC₁ receptors), in intestine of rat and PAC₁^-/- mice.

The first stated hypothesis that activation of PAC₁ receptors could have a toxic effect on the excitotoxic lesions while activation of VPAC receptors could be neuroprotective, leading to a lack of detectable effect for PACAP38, can be ruled out by the lack of protective effects of PACAP 38 in PAC₁^-/- mice [31].  In contrast, VIP neuroprotective effects are completely abolished in mice lacking VPAC₂ receptor [31]. In situ hybridization confirms the presence of VPAC₂ mRNA in the postnatal day 5 white matter [31]. When analyzed between embryonic life and adulthood, VIP specific binding site density peaks at postnatal day 5 [31]. These data suggest that, in this model, VIP-induced neuroprotection is mediated by VPAC₂ receptors. The pharmacology of this VPAC₂ receptor seems unconventional as i) PACAP does not mimic VIP effects, ii) PHI acts with a comparable potency and iii) PACAP 27 modestly inhibits the VIP specific binding while for PHI or VIP, inhibition is complete.