Endolymphatic hydrops is prevalent in the first weeks following cochlear implantation.

AIM: To explore morphological or electrophysiological evidence for the presence of endolymphatic hydrops (EH) in guinea pig cochleae in the first 3 months after cochlear implantation. METHODS: Dummy silastic electrodes were implanted atraumatically into the basal turn of scala tympani via a cochleostomy. Round window electrocochleography (ECochG) was undertaken prior to and after implantation. Animals survived for 1, 7, 28 or 72 days prior to a terminal experiment, when ECochG was repeated. The cochleae were imaged using micro-CT after post-fixing with osmium tetroxide to reveal the inner ear soft tissue structure. EH was assessed by visual inspection at a series of frequency specific places along the length of the cochlea, and the extent to which Reissner's membrane departed from its neutral position was quantified. Tissue response volumes were calculated. Using ECochG, the ratio of the summating potential to the action potential (SP/AP ratio) was calculated in response to frequencies between 2 and 32 kHz. RESULTS: There was minimal evidence of electrode trauma from cochlear implantation on micro-CT imaging. Tissue response volumes did not change over time. EH was most prevalent 7 days after surgery in implanted ears, as determined by visual inspection. Scala media areas were increased, as expected in cases of EH, over the first month after cochlear implantation. SP/AP ratios decreased immediately after surgery, but were elevated 1 and 7 days after implantation. CONCLUSIONS: EH is prevalent in the first weeks after implant surgery, even in the absence of significant electrode insertion trauma.

Cannabinoid receptor activation and elevated cyclic AMP reduce glutamate neurotoxicity.

Cannabinoid receptor activation in vivo reduces ischemic injury, a phenomenon that has not been successfully reproduced in vitro. Because cyclic adenosine monophosphate (cAMP) levels are radically elevated during ischemic reperfusion, but cannabinoid receptor activation reduces cAMP levels, we hypothesized that cannabinoids might prevent in vitro glutamate toxicity if reperfusion was simulated by cAMP supplementation after glutamate removal. Although neuronal cultures were unaffected by the single addition of either cannabinoid or dibutyryl cAMP (dbcAMP), glutamate toxicity was reduced by 20% when cannabinoid was present during glutamate exposure and either dbcAMP or forskolin was added after glutamate removal. Further studies revealed that cannabinoid receptor activation reduces glutamate toxicity by attenuating calcium influx through N- and P/Q-type calcium channels. The effect of glutamate exposure on neuronal cAMP levels was also examined. Glutamate exposure significantly reduced neuronal cAMP levels, although suppression was even greater when cannabinoid was present. Because neurological outcome after ischemia is poor when cAMP levels during reperfusion are low, it is hypothesized that cAMP elevation after glutamate exposure may offset excitotoxic and/or cannabinoid receptor-induced cAMP depletion. Cannabinoids protect against ischemic injury in vivo, but only reduce toxicity in vitro when cAMP levels are elevated, possibly suggesting that cAMP elevation during reperfusion reduces brain injury by off-setting the effect of Gi/o protein-coupled systems on adenylate cyclase.

Neuroprotective antioxidants from marijuana.

Cannabidiol and other cannabinoids were examined as neuroprotectants in rat cortical neuron cultures exposed to toxic levels of the neurotransmitter, glutamate. The psychotropic cannabinoid receptor agonist delta 9-tetrahydrocannabinol (THC) and cannabidiol, (a non-psychoactive constituent of marijuana), both reduced NMDA, AMPA and kainate receptor mediated neurotoxicities. Neuroprotection was not affected by cannabinoid receptor antagonist, indicating a (cannabinoid) receptor-independent mechanism of action. Glutamate toxicity can be reduced by antioxidants. Using cyclic voltametry and a fenton reaction based system, it was demonstrated that Cannabidiol, THC and other cannabinoids are potent antioxidants. As evidence that cannabinoids can act as an antioxidants in neuronal cultures, cannabidiol was demonstrated to reduce hydroperoxide toxicity in neurons. In a head to head trial of the abilities of various antioxidants to prevent glutamate toxicity, cannabidiol was superior to both alpha-tocopherol and ascorbate in protective capacity. Recent preliminary studies in a rat model of focal cerebral ischemia suggest that cannabidiol may be at least as effective in vivo as seen in these in vitro studies.

Cannabidiol and (-)Delta9-tetrahydrocannabinol are neuroprotective antioxidants.

The neuroprotective actions of cannabidiol and other cannabinoids were examined in rat cortical neuron cultures exposed to toxic levels of the excitatory neurotransmitter glutamate. Glutamate toxicity was reduced by both cannabidiol, a nonpsychoactive constituent of marijuana, and the psychotropic cannabinoid (-)Delta9-tetrahydrocannabinol (THC). Cannabinoids protected equally well against neurotoxicity mediated by N-methyl-D-aspartate receptors, 2-amino-3-(4-butyl-3-hydroxyisoxazol-5-yl)propionic acid receptors, or kainate receptors. N-methyl-D-aspartate receptor-induced toxicity has been shown to be calcium dependent; this study demonstrates that 2-amino-3-(4-butyl-3-hydroxyisoxazol-5-yl)propionic acid/kainate receptor-type neurotoxicity is also calcium-dependent, partly mediated by voltage sensitive calcium channels. The neuroprotection observed with cannabidiol and THC was unaffected by cannabinoid receptor antagonist, indicating it to be cannabinoid receptor independent. Previous studies have shown that glutamate toxicity may be prevented by antioxidants. Cannabidiol, THC and several synthetic cannabinoids all were demonstrated to be antioxidants by cyclic voltametry. Cannabidiol and THC also were shown to prevent hydroperoxide-induced oxidative damage as well as or better than other antioxidants in a chemical (Fenton reaction) system and neuronal cultures. Cannabidiol was more protective against glutamate neurotoxicity than either ascorbate or alpha-tocopherol, indicating it to be a potent antioxidant. These data also suggest that the naturally occurring, nonpsychotropic cannabinoid, cannabidiol, may be a potentially useful therapeutic agent for the treatment of oxidative neurological disorders such as cerebral ischemia.

Oleamide potentiates benzodiazepine-sensitive gamma-aminobutyric acid receptor activity but does not alter minimum alveolar anesthetic concentration.

UNLABELLED: A naturally occurring brain lipid, cis-9,10-octadeceamide--oleamide (OA), is found in increased concentrations in the cerebrospinal fluid of sleep-deprived cats, which suggests that it may be an endogenous sleep-inducing substance. We studied the effects of this fatty-acid derivative on the function of cloned gamma-aminobutyric acid (GABA(A)) receptors expressed in Xenopus oocytes. Oocytes were injected with cRNA synthesized in vitro to express simple GABA(A) receptors (alpha1beta1, alpha3beta1, alpha5beta1, and alpha1beta2 subunit combinations) and receptors in which the GABA-induced chloride currents were potentiated in the presence of benzodiazepines (alpha1beta1gamma2s and alpha1beta2gamma2s subunit combinations). OA only produced significant potentiation of the peak Cl- current when applied with GABA to benzodiazepine-sensitive GABA(A) receptors. The peak currents of the simple GABA(A) receptors in the presence of OA were either unaffected or slightly inhibited by OA, but the overall mean currents were not significantly altered. Oleic acid was also capable of potentiating benzodiazepine-sensitive GABA(A) receptor function. The function of other ligand-gated ion channels, such as the N-methyl-D-aspartate receptor (NR1 + NR2A or 2C) and the 5-HT3 receptor expressed in Xenopus oocytes, were unaffected by OA. Sprague-Dawley rats receiving intraperitoneal injections of oleamide (10, 20, or 100 mg/kg) showed no change in the minimum alveolar anesthetic concentration (MAC) of desflurane required to abolish movement in response to noxious (tail clamp) stimulation (control MAC 6.48% +/- 1.28% atm; 100 mg/kg OA MAC 7.05% +/- 0.42% atm). These results reinforce the view that oleyl compounds may be natural modulators of inhibitory ion channel function, but that these effects contribute little to the central nervous system depression produced by volatile anesthetics as measured by MAC. IMPLICATIONS: The putative sleep-inducing substance, oleamide, potentiates benzodiazepine-sensitive gamma-aminobutyric acid receptor function but does not alter desflurane minimum alveolar anesthetic concentration in rats.

Dual effects of anandamide on NMDA receptor-mediated responses and neurotransmission.

Anandamide is an endogenous ligand of cannabinoid receptors that induces pharmacological responses in animals similar to those of cannabinoids such as delta9-tetrahydrocannabinol (THC). Typical pharmacological effects of cannabinoids include disruption of pain, memory formation, and motor coordination, systems that all depend on NMDA receptor mediated neurotransmission. We investigated whether anandamide can influence NMDA receptor activity by examining NMDA-induced calcium flux (deltaCa2+NMDA) in rat brain slices. The presence of anandamide reduced deltaCa2+NMDA and the inhibition was disrupted by cannabinoid receptor antagonist, pertussis toxin treatment, and agatoxin (a calcium channel inhibitor). Whereas these treatments prevented anandamide inhibiting deltaCa2+NMDA, they also revealed another, underlying mechanism by which anandamide influences deltaCa2+NMDA. In the presence of cannabinoid receptor antagonist, anandamide potentiated deltaCa2+NMDA in cortical, cerebellar, and hippocampal slices. Anandamide (but not THC) also augmented NMDA-stimulated currents in Xenopus oocytes expressing cloned NMDA receptors, suggesting a capacity to directly modulate NMDA receptor activity. In a similar manner, anandamide enhanced neurotransmission across NMDA receptor-dependent synapses in hippocampus in a manner that was not mimicked by THC and was unaffected by cannabinoid receptor antagonist. These data demonstrate that anandamide can modulate NMDA receptor activity in addition to its role as a cannabinoid receptor ligand.

Anandamide hydroxylation by brain lipoxygenase:metabolite structures and potencies at the cannabinoid receptor.

Anandamide (arachidonyl ethanolamide) is a compound that was identified from porcine brain lipids by its ability to bind to the brain cannabinoid receptor. This study assessed anandamide as a substrate for a brain lipoxygenase and characterised the brain metabolite 12-hydroxyanandamide. Anandamide was also compared with arachidonic acid as a lipoxygenase substrate by examining enzyme kinetics in the presence of either of the two compounds. In addition, a non-mammalian enzyme was used to generate 11- and 15-hydroxy-anandamide in order to compare the cannabinomimetic properties of a range of anandamide derivatives. A ligand displacement assay indicated a large variation in the affinity of anandamide metabolites for the brain cannabinoid receptor. The brain metabolite, 12-hydroxyanandamide had an affinity twice that of anandamide, although the 11- and 15- hydroxy-metabolites were considerably poorer ligands of this receptor. Consistent with the receptor binding data, 12-hydroxyanandamide (unlike 15-hydroxyanandamide) inhibited forskolin-stimulated cAMP synthesis, indicating it to be a functional agonist at the brain cannabinoid receptor. Pharmacological studies of the capacity of anandamide and its metabolites to inhibit the murine vas deferens twitch response indicated the 12-hydroxy-metabolite to be less active than the parent compound, but a better cannabinomimetic than 15-hydroxyanandamide.

Biosynthesis of eicosanoids by blood cells of the crab, Carcinus maenas.

Blood cells from the crab, Carcinus maenas, stimulated with calcium ionophore A23187, in the presence of exogenous fatty acid, produced cyclooxygenase, lipoxygenase and monooxygenase derivatives of eicosatetraenoic (20:4(n - 6)) and eicosapentaenoic (20:5(n - 3)) acids. Isolation, identification and quantification of these products was achieved using chiral and reverse phase-high performance liquid chromatography, gas-chromatography, radioimmunoassay and gas chromatography-mass spectrometry. The principle metabolites observed were 8-hydroxy fatty acids and 'E' series prostaglandins. Smaller amounts of thromboxane B2, 6-keto-prostaglandin F1 alpha and 5-, 9-, 11-, 12- and 15-hydroxy-eicosatetraenoic acids were also synthesised. Lipoxygenase, cyclooxygenase and cytochrome P-450 inhibitors were used to investigate the mode of product formation. Mixtures of hydroxy-fatty acid enantiomers were produced and the dominant chiral form varied with the position of the hydroxyl group. No leukotrienes or lipoxins were detected.