histogranin and Disease-Models--Animal

The treatment of spinal cord injury (SCI)-induced neuropathic pain presents a challenging healthcare problem. The lack of available robust pharmacological treatments underscores the need for novel therapeutic methods and approaches. Due to the complex character of neuropathic pain following SCI, therapies targeting multiple mechanisms may be a better choice for obtaining sufficient long-term pain relief. Previous studies in our lab showed analgesic effects using combinations of an NMDA antagonist peptide [Ser1]histogranin (SHG), and the mu-opioid peptides endomorphins (EMs), in several pain models. As an alternative to drug therapy, this study evaluated the analgesic potential of these peptides when delivered via gene therapy.. Lentiviruses encoding SHG and EM-1 and EM-2 were intraspinally injected, either singly or in combination, into rats with clip compression SCI 2 weeks following injury. Treated animals showed significant reduction in mechanical and thermal hypersensitivity, compared to control groups injected with GFP vector only. The antinociceptive effects of individually injected components were modest, but the combination of EMs and SHG produced robust and sustained antinociception. The onset of the analgesic effects was observed between 1-5 weeks post-injection and sustained without decrement for at least 7 weeks. No adverse effects on locomotor function were observed. The involvement of SHG and EMs in the observed antinociception was confirmed by pharmacologic inhibition using intrathecal injection of either the opioid antagonist naloxone or an anti-SHG antibody. Immunohistochemical analysis showed the presence of SHG and EMs in the spinal cord of treated animals, and immunodot-blot analysis of CSF confirmed the presence of these peptides in injected animals. In a separate group of rats, delayed injection of viral vectors was performed in order to mimic a more likely clinical scenario. Comparable and sustained antinociceptive effects were observed in these animals using the SHG-EMs combination vectors compared to the group with early intervention.. Findings from this study support the potential for direct gene therapy to provide a robust and sustained alleviation of chronic neuropathic pain following SCI. The combination strategy utilizing potent mu-opioid peptides with a naturally-derived NMDA antagonist may produce additive or synergistic analgesic effects without the tolerance development for long-term management of persistent pain.

Topics: Animals; Cell Line, Tumor; Disease Models, Animal; Genetic Vectors; Humans; Hyperalgesia; Lentivirus; Male; Neuralgia; Neuroblastoma; Neuropeptides; Opioid Peptides; Pain Measurement; Pain Threshold; Proteins; Rats; Rats, Sprague-Dawley; Recovery of Function; Spinal Cord Injuries

Excitatory amino acids (EAAs) play an important role during ischemic brain injury. In this study we examined the protective effect of histogranin (HN), an endogenous peptide that antagonizes excitatory amino acids-mediated activity noncompetitively, in an animal model of cerebral ischemia. Adult rats were anesthetized with chloral hydrate. Histogranin was given intracerebroventricularly before a 60-min middle cerebral artery occlusion (MCAo). Animals were examined for their locomotor activity 2 days after MCAo. Histogranin significantly increased locomotor activity in the stroke rats. Histogranin pretreatment reduced the volume of cerebral infarction and the caspase-3 immunoreactivity in the stroke animals. Taken together, our data suggest that histogranin is protective against ischemic brain injury. The protective effect may involve anti-apoptotic mechanisms.

Topics: Analysis of Variance; Animals; Behavior, Animal; Brain Injuries; Caspase 3; Caspases; Disease Models, Animal; Excitatory Amino Acid Antagonists; Functional Laterality; Ischemic Attack, Transient; Male; Motor Activity; Proteins; Rats; Rats, Sprague-Dawley; Tetrazolium Salts

Histogranin (HN)-like nonpeptides were designed and synthesized using benzimidazole (compound 1) and o-phenylenediamine (compounds 2-7) as scaffolds for the attachment of phenolic hydroxyl and basic guanidino pharmacophoric elements present in HN. The benzimidazole derivative N-5-guanidinopentanamide-(2R)-yl-2-(p-hydroxybenzyl)-5-carboxybenzimidazole (1) and the o-phenylenediamine derivative N-5-guanidinopentanamide-(2S)-yl-2-N-(p-hydroxyphenylacetyl) phenylenediamine (2) were more potent analgesics than HN in both the mouse writhing (5.5 and 3.5 as potent as HN, respectively) and tail-flick (11.8 and 8.0 as potent as HN, respectively) pain assays. Improvements in the potencies and times of action of compound 2 in the mouse writhing test were obtained by attaching carboxyl (6)or p-Cl-benzoyl (7) groups at position 4 of the (2R) o-phenylenediamine derivative (5). In rats, compounds 2 (80 nmol i.t.), 6 (36 nmol i.t.), and 7 (18 nmol i.t.) were effective in blocking both persistent inflammatory pain in the formalin test and hyperalgesia in the complete Freund adjuvant assay. Compounds 2, 6, and 7, but not compound 1 at 10 nmol (i.c.v.) also mimicked the HN (60 nmol i.c.v.) blockade of N-methyl-D-aspartate (NMDA)-induced convulsions in mice. Finally, in primary cultures of rat alveolar macrophages, HN and compounds 1, 2, 6, and 7 (10(-8) M) significantly blocked lipopolysaccharide-induced cyclooxygenase-2 induction and prostaglandin E(2) secretion. These studies indicate that both derivatives of benzimidazole and o-phenylenediamine mimic the in vivo antinociceptive and in vitro anti-inflammatory effects of HN, but the HN protection of mice against NMDA-induced convulsions is mimicked only by the o-phenylenediamine derivatives.

Topics: Analgesics; Animals; Anti-Inflammatory Agents; Benzimidazoles; Cyclooxygenase 2; Dinoprostone; Disease Models, Animal; Drug Interactions; Isoenzymes; Lipopolysaccharides; Macrophages; Male; Mice; Pain; Phenylenediamines; Prostaglandin-Endoperoxide Synthases; Proteins; Rats; Rats, Sprague-Dawley