Endothelium-Mediated Action of Analogues of the Endogenous Neuropeptide Kyotorphin (Tyrosil-Arginine): Mechanistic Insights from Permeation and Effects on Microcirculation

Kyotorphin (KTP) is an endogenous peptide with analgesic properties when administered into the central nervous system (CNS). Its amidated form (L-Tyr-L-Arg-NH2; KTP-NH2) has improved analgesic efficacy after systemic administration, suggesting blood-brain barrier (BBB) crossing. KTP-NH2 also has anti-inflammatory action impacting on microcirculation. In this work, selected derivatives of KTP-NH2 were synthesized to improve lipophilicity and resistance to enzymatic degradation while introducing only minor changes in the chemical structure: N-terminal methylation and/or use of D amino acid residues. Intravital microscopy data show that KTP-NH2 having a D-Tyr residue, KTP-NH2-DL, efficiently decreases the number of leukocyte rolling in a murine model of inflammation induced by bacterial lipopolysaccharide (LPS): down to 46% after 30 min with 96 mu M KTP-NH2-DL. The same molecule has lower ability to permeate membranes (relative permeability of 0.38) and no significant activity in a behavioral test which evaluates thermal nociception (hot-plate test). On the contrary, methylated isomers at 96 mu M increase leukocyte rolling up to nearly 5-fold after 30 min, suggesting a proinflammatory activity. They have maximal ability to permeate membranes (relative permeability of 0.8) and induce long-lasting antinociception

Inhibition of nociceptive responses after systemic administration of amidated kyotorphin.

BACKGROUND AND PURPOSE: Kyotorphin (KTP; L-Tyr-L-Arg), an endogenous neuropeptide, is potently analgesic when delivered directly to the central nervous system. Its weak analgesic effects after systemic administration have been explained by inability to cross the blood-brain barrier (BBB) and detract from the possible clinical use of KTP as an analgesic. In this study, we aimed to increase the lipophilicity of KTP by amidation and to evaluate the analgesic efficacy of a new KTP derivative (KTP-amide - KTP-NH(2) ). EXPERIMENTAL APPROACH: We synthesized KTP-NH(2) . This peptide was given systemically to assess its ability to cross the BBB. A wide range of pain models, including acute, sustained and chronic inflammatory and neuropathic pain, were used to characterize analgesic efficacies of KTP-NH(2) . Binding to opioid receptors and toxicity were also measured. KEY RESULTS: KTP-NH(2) , unlike its precursor KTP, was lipophilic and highly analgesic following systemic administration in several acute and chronic pain models, without inducing toxic effects or affecting motor responses and blood pressure. Binding to opioid receptors was minimal. KTP-NH(2) inhibited nociceptive responses of spinal neurons. Its analgesic effects were prevented by intrathecal or i.p. administration of naloxone. CONCLUSIONS AND IMPLICATIONS: Amidation allowed KTP to show good analgesic ability after systemic delivery in acute and chronic pain models. The indirect opioid-mediated actions of KTP-NH(2) may explain why this compound retained its analgesic effects although the usual side effects of opioids were absent, which is a desired feature in next-generation pain medications.

High resolution Raman and neutron investigation of Mg(BH4)2 in an extensive temperature range.

Raman spectra of Mg(BH(4))(2) have been measured in an extensive temperature range, from 15 to 473 K. Taking into account the high temperature conversion from the alpha to the beta phase, we have observed evident signatures of this phase transition and determined the Raman vibrational spectrum of each phase. The neutron scattering spectra of the beta phase sample were also recorded. The present experimental results have been compared to the density functional theory calculations available in the literature, and a substantial agreement has been found.

Synthesis and characterization of NaBD3H, a potential structural probe for hydrogen storage materials.

Specifically labeled NaBD(3)H has been synthesized and characterized using X-ray diffraction, NMR, and vibrational spectroscopy. The isotopic purity of the compound, as estimated from NMR spectra, was found to be about 85% with the compound NaBD(2)H(2) as the second product. IR spectra confirm the relatively strong intensity of the single B-H stretching mode predicted from DFT calculations. Anharmonic DFT calculations show that for the BD(3)H(-) ion Fermi resonances with the single B-H stretching mode are very limited, making this mode a promising structural probe for complex borohydrides which can be prepared by metathetical reactions.

Differential effects of Mg2+ and other divalent cations on the binding of tritiated opioid ligands.

The effects of MgCl2 on the binding of tritiated ligands to opioid binding sites in homogenates of guinea-pig brain in HEPES buffer have been studied. The binding of tritiated mu-, delta-, and kappa-opioid agonists was promoted in a concentration-dependent manner over a range of MgCl2 concentrations from 0.1 mM to 10 mM, as was binding of the nonselective antagonists [3H]diprenorphine and [3H]naloxone. At concentrations of MgCl2 above 10 mM reversal of this effect was observed. The effects of MgCl2 on binding parameters differed at each site. The promoting effects of MgCl2 were mimicked by MnCl2, CaCl2, and MgSO4, but CoCl2 and ZnCl2 were inhibitory. Following treatment of guinea-pig brain synaptosomes at pH 11.5 to eliminate G proteins, the binding of the mu-opioid agonist [3H][D-Ala2, MePhe4, Gly-ol5]enkephalin and [3H]naloxone was much reduced but binding of [3H]diprenorphine was unaffected. Under these conditions MgCl2 still promoted binding of [3H]diprenorphine. The results suggest that Mg2+ ions promote binding by an action at the opioid receptor, even in the absence of G protein, and that opioid antagonists may differ in their recognition of opioid receptor binding sites.