Chemical excitation and inactivation in photoreceptors of the fly mutants trp and nss.

The Drosophila and Lucilia photoreceptor mutants, trp and nss, respond like wild-type flies to a short pulse of intense light or prolonged dim light; however, upon continuous intense illumination, the trp and nss mutants are unable to maintain persistent excitation. This defect manifests itself by a decline of the receptor potential toward baseline during prolonged intense illumination with little change in the shape or amplitude of the quantal responses to single photons (quantum bumps). Previous work on the trp and nss mutants suggests that a negative feedback loop may control the rate of bump production. Chemical agents affecting different steps of the phototransduction cascade were used in conjunction with light to identify a possible branching point of the feedback loop and molecular stages which are affected by the mutation. Fluoride ions, which in the dark both excite and adapt the photoreceptors of wild-type flies, neither excite nor adapt the photoreceptors of the trp and nss mutants. The hydrolysis-resistant analogue, GTP gamma S, which excites the photoreceptors of wild-type flies, resulting in noisy depolarization, markedly reduces the light response of both mutant flies. Intracellular recordings revealed, however, that the inhibitory effect of GTP gamma S on the nss mutant was accompanied neither by any significant depolarization nor by an increase in the noise, and thus was very different from the effect of a dim background light. The combination of inositol trisphosphate and diphosphoglycerate (InsP3 + DPG), which efficiently excites the photoreceptors of wild-type Lucilia, also excites the photoreceptors of nss Lucilia mutant. The InsP3 + DPG together act synergistically with light to accelerate the decline of the response to light in the mutant flies. These results suggest that the fly phototransduction pathway involves a feedback regulatory loop, which branches subsequent to InsP3 production and regulates guanine nucleotide-binding protein (G protein)-phospholipase C activity. A defect in this regulatory loop, which may cause an unusually low level of intracellular Ca2+, severely reduces the triggering of bumps in the mutants during intense prolonged illumination.

Comparative antimicrobial activity of the new macrolides against Borrelia burgdorferi.

The in vitro and in vivo activity of the new macrolides azithromycin, clarithromycin and roxythromycin was compared with that of erythromycin against Borrelia burgdorferi. In in vitro tests using ten clinical isolates all macrolides were highly active against Borrelia burgdorferi (MIC90 0.015-0.06 micrograms/ml). Azithromycin was more potent than the other macrolides in experimental animal infection, eradicating the organism in all animals tested at a dosage of 8 mg/kg.

[Allergic reactions of the immediate and delayed type following prednisolone medication]. / Allergische Reaktionen vom Sofort- und Spättyp nach Prednisolon-Medikation.

Allergic sensitization due to corticosteroids seldom occur and are compound-specific, as a rule. They induce various clinical features, in particular urticaria, different exanthematous reactions and contact dermatitis as shown by our observations in 10 patients with prednisolone-allergy. In three cases a simultaneous allergy due to methylprednisolone was found. Impairment of the features or alterations in the clinical morphology after the application of prednisolone suggest the diagnosis, which can be confirmed by means of the scratch and epicutaneous testing and in one patient by the oral exposure, additionally. In addition a delayed-type sensitization to propylene glycol could be proved in three cases, and the same was with romulgin and parabens in one case each. Dexamethasone was used and tolerated as alternative and emergency medication in equivalent doses.

Light reduces the excitation efficiency in the nss mutant of the sheep blowfly Lucilia.

The nss (no steady state) phototransduction mutant of the sheep blowfly Lucilia was studied electrophysiologically using intracellular recordings. The effects of the nss mutation on the receptor potential are manifested in the following features of the light response. (a) The responses to a flash or to dim lights are close to normal, but the receptor potential decays close to the baseline level during prolonged illumination after a critical level of light intensity is reached. (b) The decline of the response is accompanied by a large reduction in responsiveness to light that recovers within 20 s in the dark. (c) The full reduction in responsiveness to light is reached when approximately 13% of the photopigment molecules are converted from rhodopsin (R) to metarhodopsin (M). (d) A maximal net pigment conversion from R to M by blue light induces persistent inactivation in the dark, without an apparent voltage response. This inactivation could be abolished at any time by M-to-R conversion with orange light. The above features of the mutant indicate that the effect of the nss mutation on the light response of Lucilia is very similar to the effects of the transient receptor potential (trp) mutation on the photoreceptor potential of Drosophila. Noise analysis and voltage measurements indicate that the decay of the receptor potential is due to a severe reduction in the rate of occurrence of the elementary voltage responses (bumps). The bumps are only slightly modified in shape and amplitude during the decline of the response to light of medium intensity. There is also a large increase in response latency during intense background illumination. These results are consistent with the hypothesis that separate, independent mechanisms determine bump triggering and bump shape and amplitude. The nss mutation affects the triggering mechanism of the bump.

Coupling of photoexcited rhodopsin to inositol phospholipid hydrolysis in fly photoreceptors.

Fly photoreceptor membranes were used to test the effect on defined biochemical reactions of light and of compounds causing photoreceptor excitation. Complementary electrophysiological studies examined whether putative second messengers excite the fly photoreceptor cells. This analysis revealed the following sequence of events: photoexcited rhodopsin activates a G protein by facilitating GTP binding. The G protein then activates a phospholipase C that generates inositol trisphosphate, which in turn acts as an internal messenger to bring about depolarization of the photoreceptor cell. Binding assays of GTP analogs and measurements of GTPase activity showed that there are 1.6 million copies of G protein per photoreceptor cell. The GTP binding component is a 41-kDa protein, and the light-activated GTPase is dependent on photoconversion of rhodopsin to metarhodopsin. Analysis of phospholipase C activity revealed that this enzyme is under stringent control of the G protein, that the major product formed is inositol trisphosphate, and that this product is rapidly hydrolyzed by a specific phosphomonoesterase. Introduction of inositol trisphosphate to the intact photoreceptor cell mimics the effect of light, and bisphosphoglycerate, which inhibits inositol trisphosphate hydrolysis, enhances the effects of inositol trisphosphate and of dim light. The interaction of photoexcited rhodopsin with a G protein is thus similar in both vertebrate and invertebrate photoreceptors. These G proteins, however, activate different photoreceptor enzymes: phospholipase C in invertebrates and cGMP phosphodiesterase in vertebrates.

In vitro and in vivo susceptibility of Borrelia burgdorferi.

The antispirochetal activity in vitro and in vivo of several antibiotics against ten isolates of Borrelia burgdorferi from human spinal fluids and skin biopsies was determined. Borrelia burgdorferi was most susceptible in vitro to erythromycin, ceftriaxone and cefotaxime (MIC90: 0.06, 0.06, 0.12 mcg/ml respectively). Less activity was observed with tetracycline, amoxycillin and lincomycin (MIC90: 0.50 mcg/ml), imipenem and augmentin (MIC90: 0.25 mcg/ml), oxacillin (MIC90: 1 mcg/ml), ciprofloxacin (MIC90: 2 mcg/ml) and ofloxacin (MIC90: 4 mcg/ml). Penicillin G, normally regarded as appropriate treatment for Lyme disease, had an MIC90 of only 4 mcg/ml. With the exception of erythromycin, activity in vitro corresponded to the activity in vivo. Erythromycin, however, was less active in vivo, and penicillin G showed poor activity both in vitro and in vivo.

[Methodical problems in detecting new allergens in ;animal experiments (author's transl)]. / Methodische Probleme der tierexperimentellen Erfassung neuer Allergene.

With the help of the Tina-test partial aspects of the predictive testing of potential allergens were examined in animal experiments. The Tina-test is based on the intramuscular, intracutaneous and epicutaneous application of the allergen in question using Freund's adjuvant and sodium laurylsulphate. The test has been acknowledged by the authorities as a standardized method. the following aspects were examined. A prolongation of the exposition times causes an increase of the sensitization rates. Freund's adjuvant should be used in every case, since the percentage of sensitized animals may be clearly increased. Outbred guinea pigs are well suited. the use of inbred animals is not necessary. In cases where more than 30% of the experimental group fall ill during the procedure of sensitization, the results are not reliable. Neither the sex of the animals nor the seasons influence sensitization rates. Per experimental group at least 25 animals should be used. Pilot studies with different quantities of allergens (potassium bichromate: 131 mg: 13.1 mg and 1.31 mg) did not show a clear influence on the results. Similar results we received also with nickel-2-sulphate. The sensitization rate corresponded well with our clinical experience in 19 out of 24 substances. With turpentine, nickel, mercury and a low-molecular phenol resin the number of the sensitized animals was somewhat too low, whereas with tetramethylthiuramdisulfide the percentage was somewhat too high.

[The allergenic effect of rubber accelerators tetramethyl thiuram disulfide (TMTD) and mercaptobenzothiazole (MBT)]. / Die allergene Wirkung der Gummiakzeleratoren Tetramethylthiuramdisulfid (TMTD) und Mercaptobenzothiazol (MBT)

The eczematogenic effects of tetramethylthiuramidsulfid and of mercaptobenzothiazol were studied on the basis of an inquiry in establishments using TMTD and MBT, a frequency analysis of positive reactions of our standard series in 1971 and testing of various rubber articles on sensitized persons as well as animal experiments. The results indicate a wider use, a higher percentage of positive epicutaneous tests and also a higher sensitizing potency in the animal test of the TMTD as compared with MBT. MBT was removed from our standard series.