Hypocrellin A (HA), a peryloquinone derivative, has recently been isolated from a fungus Hypocrella bambusae. This lipid soluble pigment, in combination with phototherapy, has been used to treat many skin diseases including the keloids caused by scalding and burns. We have studied the effects of photosensitized HA on biomembranes using pig heart microsomes. Photosensitization of HA was found to peroxidize the membrane lipids in the cardiac microsomes. The photodamage imposed by HA depended not only on the concentration of HA but also on the time of irradiation and pH of the system. Superoxide dismutase (SOD), ascorbic acid, beta-carotene and 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) inhibited the lipid peroxidation approximately 50, approximately 50, approximately 30 and approximately 97%, respectively. Spin trapping in combination with EPR spectroscopic techniques was used to identify the reactive free radicals during the photoreaction. Formation of superoxide anion radical, (O2-.), was identified by the SOD-inhibitable DMPO-O2- EPR spectrum. Both SOD and ascorbic acid inhibited the EPR signal intensity in a dose-dependent manner with rate constants of 6.78 x 10(8) M-1 s-1 and 1.82 x 10(4) M-1 s-1, respectively. The lifetime of O2-., under these conditions, was found to be 1.1 s. Photoirradiation of HA yielded a HA free radical with a g = 2.002 which was not suppressed by SOD but in the presence of reductants such as ascorbic acid and catechol the septum was completely suppressed. The increase of the EPR signal intensity and malondialdehyde formation with increasing pH may be due, in part, to the production of predominant *HA- species at high pH which would be more reactive with oxygen to yield O2-.. These results indicate that the lipid peroxidation of the cardiac membranes observed during photooxidation of HA may arise, in part, from the interaction of membrane lipids with reactive species of oxygen and HA free radical produced during the photo-irradiation.

We have previously demonstrated increases in serotonin (5-HT) content and immunoreactivity within spinal trigeminal subnucleus interpolaris (SpVi) that are correlated with the functional changes observed in this subnucleus following adult infraorbital nerve (ION) transection. To assess the possible functional significance of this change, we have examined the influence of 5-HT afference upon the normal response properties of cells in SpVi. We employed local depletion of the transmitter, using 5,7-dihydroxtryptamine (5,7-DHT), in combination with extracellular single-cell recording. Chromatographic methods revealed a 97.6% depletion of 5-HT 24 hr after neurotoxin injection. Immunocytochemical procedures revealed depletion of 5-HT throughout SpVi. Physiological recordings were made from 403 SpVi cells in 5,7-DHT-injected rats and 387 cells in vehicle-injected rats. All recordings were made 19-27 hr after injection. Horseradish peroxidase (HRP) deposits from the recording electrode were used to mark recording tracks. 5-HT depletion did not influence receptive field (RF) location, size, or continuity, or the dynamic response characteristics of SpVi cells. It did, however, (1) alter the probability that certain types of somatosensory receptor surfaces would activate local-circuit neurons, and (2) influence the rate of firing of spontaneously active SpVi cells. There was a significant increase in the proportion of vibrissa-sensitive cells with infraorbital RF components, and a concurrent decrease in the proportion of guard-hair-sensitive cells. It therefore appears that 5-HT input to SpVi is necessary for some mechanoreceptive features of the normal functional organization of this area. These functional changes were interesting in that they were opposite to those found following adult ION transection, which increases 5-HT within SpVi. Thus, changes in 5-HT central afference to SpVi that follow ION damage may be responsible for at least one type of functional change observed following this peripheral lesion.

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