Endomorphin 1 and endomorphin 2 suppress in vitro antibody formation at ultra-low concentrations: anti-peptide antibodies but not opioid antagonists block the activity.

Endomorphin 1 (EM-1) and endomorphin 2 (EM-2) were tested for their capacity to alter immune function. Addition of either of these peptides to murine spleen cells in vitro inhibited antibody formation to sheep red blood cells in a bi-phasic dose dependent manner. Maximal inhibition was achieved at doses in the range of 10(-13) to 10(-15)M. Neither naloxone (general opioid receptor antagonist) nor CTAP (selective mu opioid receptor antagonist) blocked the immunosuppressive effect. To show that there was specificity to the immunosuppressive activity of the peptides, affinity-purified rabbit antibodies were raised against each of the synthetic EM peptides haptenized to KLH and tested for capacity to inhibit immunosuppression. Antibody responses were monitored by a standard solid phase antibody capture ELISA, and antibodies were purified by immunochromatography using the synthetic peptides coupled to a Sepharose 6B resin. Verification of the specificity of affinity-purified antisera was performed by immunodot-blot and solid-phase RIA assays. The antisera specific for both EM-1 and EM-2 neutralized the immunosuppressive effects of their respective peptides in a dose-related manner. Control normal rabbit IgG had no blocking activity on either EM-1 or EM-2. These studies show that the endomorphins are immunomodulatory at ultra-low concentrations, but the data do not support a mechanism involving the mu-opioid receptor.

Orphanin-FQ/nociceptin inhibits kindling epileptogenesis and enhances hippocampal feed-forward inhibition.

The role of Orphanin-FQ/nociceptin in synaptic plasticity was assessed by its potency in modulating kindling epileptogenesis in vivo, and feed-forward inhibition in hippocampal recordings in vitro. In addition, a specific rabbit antiserum against this peptide was obtained and the immunohistochemical distribution of nociceptin was determined in rat brain slices. After the establishment of kindling epilepsy, by daily electrical stimulation of the piriform cortex, the i.c.v. injection of nociceptin, 20 min before the kindling stimulation, was not able to block the generation of the generalized seizures, nor to alter their duration. However, the i.c.v. injection of nociceptin, 20 min before each stimulation along the kindling process, depressed its development in a dose-dependent manner. This effect was specific since the nociceptin antagonist [Phe1psi(CH2-NH)Gly2]NC(1-13)NH2, but not the broad-spectrum opiate antagonist, naloxone, was able to completely block nociceptin actions. The inhibitory role of nociceptin was assessed by in vitro recordings from entorhinal cortex-hippocampal slices. By single pulses applied over the Schaffer collaterals, we found that synaptic transmission was facilitated onto CA1, but using a paired-pulse protocol, we found that nociceptin potentiated feed-forward inhibition. The immunohistochemical data show that nociceptin is expressed in limbic cortical regions, including the piriform cortex and the hippocampus. Our results demonstrate that nociceptin exerts a modulatory role in limbic excitability and suggest that it provides an inhibitory control in the development of epilepsy by possibly inhibiting the spread of excitation through the system, by favoring feed-forward inhibition.