Evidence that the agonist action of dynorphin A(1-8) in the guinea-pig myenteric-plexus may be mediated partly through conversion to [Leu5]enkephalin.

1. The agonist action of the opioid peptide dynorphin A(1-8) on the myenteric plexus-longitudinal muscle of the guinea-pig ileum has been characterized. 2. The endogenous opioid peptide dynorphin A(1-8) was rapidly degraded by slices of myenteric plexus-longitudinal muscle of the guinea-pig ileum. 3. A product of the degradation was the delta-receptor preferring [Leu5]enkephalin. Levels of [Leu5]enkephalin were markedly increased in the presence of the peptidase inhibitors bestatin, thiorphan and captopril. 4. In the myenteric plexus dynorphin A(1-8) acted as a kappa-receptor agonist. In the presence of bestatin, thiorphan and captopril a mu-receptor agonist effect was observed. This mu-agonist action was lost in the presence of N-[1-(RS)-carboxy-2-phenylethyl]Ala-Ala-Phe-p-aminobenzoate, an inhibitor of the endopeptidase enzyme EC 3.4.24.15. 5. The results suggest that formation of [Leu5]enkephalin from dynorphin A(1-8) may be an important conversion process. The enzyme responsible may be the Zn2(+)-metalloendopeptidase, EC 3.4.24.15.

Conversion of dynorphin A(1-8) to [Leu5]-enkephalin in rat central nervous tissue during development.

Rat central nervous tissue contains enzymic activity that is able to convert the kappa-receptor preferring opioid peptide dynorphin A(1-8) to the delta-nu-receptor preferring opioid peptide [Leu5]enkephalin. The ontogeny of this conversion process has been studied in vitro using cortex, striatum, cerebellum and spinal cord tissues of the developing rat brain. Evidence for the enzymic cleavage of the Leu5-Arg6 bond of dynorphin A(1-8) to afford [Leu5]enkephalin was observed as early as neonatal day 1. The degree of conversion increased up to day 7, at which time adult levels were attained. Results in all tissues studied were similar. The relationship between the increase in the conversion process with age and the ontogeny of opioid peptides and their receptors may indicate an important role for the process in the developing nervous system.

Acute and chronic effects of intrathecal galanin on behavioural and biochemical markers of spinal motor function in adult rats.

The spinal motor effects of galanin, which co-exists with 5-hydroxytryptamine (5-HT) and thyrotrophin-releasing hormone (TRH) in bulbospinal raphe neurones innervating spinal motoneurones, were examined by administering this neuropeptide through indwelling intrathecal cannulae to conscious adult Wistar rats. The acute effect of intrathecal galanin on spontaneous motor behaviour and the motor behaviours (back muscle contractions and wet-dog shakes) elicited by intrathecal injection of the non-selective 5-HT receptor agonist, 5-methoxy-N, N'-dimethyltryptamine (5-MeODMT) or the TRH analogue, RX 77368 analogue, RX 77368 (pGlu-His-3,3'-dimethyl-ProNH2), respectively, and the chronic effect of galanin on neurochemical markers for bulbospinal raphe neurones and spinal motoneurones were determined. Intrathecal galanin (0.1 to 10 micrograms) did not produce any notable motor behaviours when given alone, but pretreatment with the neuropeptide (0.1 micrograms) significantly attenuated both the number of wet-dog shakes and the amount of forepaw-licking induced by RX 77368, without affecting 5-MeODMT-induced back muscle contractions. Repeated intrathecal galanin administration (1 microgram, twice daily for 5 d) significantly elevated 5-HT (but not 5-hydroxyindoleacetic acid) and substance P-like immunoreactive (LI) levels and choline acetyltransferase (ChAT) activity in the dorsal, but not in the ventral, portion of the thoraco-lumbar spinal cord. In contrast, chronic intrathecal galanin did not alter the TRH- or calcitonin gene-related peptide (CGRP)-LI levels in either spinal cord region.(ABSTRACT TRUNCATED AT 250 WORDS)

Fungal flora from diabetic and non-diabetic human donor corneas.

Corneal tissue obtained at the Maryland Medical Examiners' Laboratory (Baltimore) from 47 cadaver donors was examined for the presence of potentially pathogenic fungi. Fifty-five percent of the donor corneas were positive for fungi consisting predominantly of Cladosporium spp. (11 isolates), Penicillium spp. (nine isolates), Rhodotorula spp. (six isolates), and Candida parapsilosis (four isolates). Exophiala jeanselmei (two isolates) and Wangiella dermatitidis were cultured from apparently healthy, normal corneas. Fifty-four percent of the fungi isolated from diabetic corneas were yeasts, as compared to 35% from the nondiabetic corneas. These fungi represent either normal fungal flora or opportunistic fungal pathogens, with the possible exceptions of E. jeanselmei and W. dermatitidis.

Human infectious corneal ulcer caused by Pythium insidiosum.

Pythium insidiosum is a fungus-like organism known to infect a variety of animals. In humans, the few known cases involving Pythium have included arterial infections and cellulitis. We present what we believe to be the first case of P. insidiosum recovered from a human corneal ulcer. The organism is difficult to isolate, causing delays in diagnosis. It is also resistant to the usual antifungal medications, making surgical excision the treatment of choice.

The physician healer: ancient magic or modern science?

The therapeutic role of general practitioners (GPs) is one that, over the years, has slowly diminished with the growing fashion for evidence-based medicine. However, it is clear that the art of healing and the strength of the doctor-patient relationship play a vital role in improving the well-being of patients. This is exemplified by the placebo effect, where the attitude of the doctor can make an appreciable difference to the psychological response of the patient who feels the need to be understood and listened to empathically. By maximizing the role of the physician healer, there is considerable scope for bridging the gap left by the impersonality of medical science, while at the same time increasing the GP's effectiveness.

Fungal infections: a growing threat.

THE EMERGENCE OF newly identified fungal pathogens and the reemergence of previously uncommon fungal diseases is primarily related to increases in the numbers of susceptible persons: people with HIV infection, bone marrow and organ transplant recipients, cancer patients being treated with chemotherapy, critically ill persons, and very low birth weight ( < or = 1500 g) infants. These immunocompromised populations are at risk for infection not only with opportunistic pathogens (for example, Pneumocystis, Candida, Cryptococcus, Trichosporon, Malassezia, Aspergillus, Penicillium marneffei, and numerous other moulds or yeasts) but also with fungal pathogens that usually infect otherwise healthy persons not previously exposed to endemic fungi (for example, Coccidioides immitis, Histoplasma capsulatum, and Blastomyces dermatitidis) and Sporothrix schenckii. Morbidity, mortality, and health care costs associated with fungal infections are high. Addressing the emergence of fungal diseases will require increased surveillance coupled with the availability of rapid, noninvasive diagnostic tests; monitoring the development of resistance to antifungal agents; and research focused on the understanding, prevention, and control of fungal infections.

A comparison of gait with solid, dynamic, and no ankle-foot orthoses in children with spastic cerebral palsy.

BACKGROUND AND PURPOSE: This study compared the effects of dynamic ankle-foot orthoses (DAFOs) with a plantar-flexion stop, polypropylene solid ankle-foot orthoses (AFOs), and no AFOs on the gait of children with cerebral palsy (CP). These orthoses were used to reduce the excessive ankle plantar flexion during the stance phase of gait. SUBJECTS AND METHODS: Ten children with spastic CP (6 with diplegia and 4 with hemiplegia) were tested after wearing no AFOs for an initial 2-week period, solid AFOs for 1 month, no AFOs for an additional 2 weeks, and DAFOs for 1 month. The effects of the two orthoses and no AFOs on lower-extremity muscle timing, joint motions, and temporal-distance characteristics were compared. RESULTS: Both orthoses increased stride length, decreased cadence, and reduced excessive ankle plantar flexion when compared with no orthoses. No differences were found for the gait variables when comparing the two orthoses. CONCLUSION AND DISCUSSION: Based on the data, the authors believe that although both orthoses would be recommended for children with spastic CP and excessive ankle plantar flexion during stance, additional individual factors should be considered when selecting either orthosis.

In vivo models: evaluating antifungal agents.

A summary is presented on some basic aspects of pure and applied studies involving in vivo antifungal models. A standard mouse model is described for the purpose of establishing acute, systemic fungal infection for drug testing. Three representative mycoses, candidiasis, cryptococcosis, and aspergillosis, and three representative drugs, amphotericin B (Ampho B), 5-fluorocytosine (5-FC) and ketoconazole (KTZ) are given as examples. Second, a quantitative spleen culture technique is described for obtaining quantitative data for the in vivo activity of Ampho B, KTZ and fluconazole in murine histoplasmosis. And finally, special application of the acute infection model will be made to evaluate all four of the above drugs in systemic phaeohyphomycosis with central nervous system involvement.

US-Japan workshops in medical mycology: past, present and future.

The Extramural Mycology Program of the National Institutes of Health (NIH), National Institute of Allergy and Infectious Diseases (NIAID) has organized and implemented a five workshop series in medical mycology during a critical period in the evolution of contemporary medical mycology (1992 to 2000; http://www.niaid.nih.gov/research/dmid.htm). The goals of the workshop series were to: initiate interactions; build collaborations; identify research needs; turn needs into opportunities; stimulate molecular research in medical mycology; and summarize recommendations emerging from the workshop proceedings. A recurring recommendation in the series was to foster communications within and beyond the field of medical mycology. US-Japan interactions were noted as one specific example of potential information exchange for mutual benefit. The first formal action directed at this recommendation was the workshop Emergence and Recognition of Fungal Diseases convened under the auspices of the US-Japan Cooperative Medical Science Program (USJCMSP; http://www.niaid. nih.gov/dmid/us%5Fjapan/default.htm) in Bethesda, Maryland USA on 30 June 1999 (D.M. Dixon & T. Matsumoto, co-chairs). A major goal of the workshop was to present contemporary medical mycology to the Joint Committee of the USJCMSP through representative research presentations in order to make the Committee aware of current status in the field, and the potential for scientific interactions. The second formal action is the workshop, under the auspices of the Japanese Society for Medical Mycology Medical Perspectives of Fungal Genome Studies scheduled for 28 November 2000 in Tokyo, Japan (T. Matsumoto & D.M. Dixon, co-chairs). The NIAID Mycology Workshop series recommended interactions between the following groups: academic and pharmaceutical; medical and molecular (model systems); medical and plant pathogens; basic and clinical; mycologists and immunologists. The first two US-Japan workshops can be viewed as consistent with these recommendations, and serve as a Western/Eastern gateway for exchange. The focus of the second US-Japan workshop on genome projects for the medically important fungi provides an excellent model for international communications. Given the tsunami of information that is flowing from genomics and bioinformatics, it is clear that global interactions will be essential in managing and interpreting the data.

In vitro and in vivo drug studies with three agents of central nervous system phaeohyphomycosis.

Amphotericin B (Amph B), 5-fluorocytosine (5-FC), ketoconazole (KTZ), fluconazole (FLZ), amorolfine (AMOR) and terbinafine (TER) were tested against 3 agents of central nervous system phaeohyphomycosis in vitro and in life-threatening infections in mice. The fungi studied were Cladosporium bantianum, Dactylaria constricta and Wangiella dermatitidis. The broadest protection against this group of fungi in mice was offered by 5-FC followed by Amph B and FLZ, then KTZ. AMOR and TER were inactive in vivo. The results of in vitro susceptibility testing had no predictive value. In contrast, the data obtained from the mouse models should be useful clinically.

Formation of [Leu5]enkephalin from dynorphin A(1-8) by rat central nervous tissue in vitro.

[3H]Dynorphin A(1-8) is readily metabolised by rat lumbosacral spinal cord tissue in vitro, affording a variety of products including a significant amount (20% recovered activity) of [3H][Leu5]enkephalin. In the presence of the peptidase inhibitors bestatin, captopril, thiorphan, and leucyl-leucine, [3H][Leu5]enkephalin was the major metabolic product, accounting for 60% of recovered activity. Production of [3H][Leu5]enkephalin was seen across all gross brain regions. The enzyme responsible for the cleavage has an optimal substrate length of 8-13 amino acids and is inhibited by N-[1-(RS)-carboxy-2-phenylethyl]-Ala-Ala-Phe-p-aminobenzoate, a site-directed inhibitor of the metalloendopeptidase EC 3.4.24.15. However the enzymic breakdown also has properties in common with involvement of endo-oligopeptidase A. Possible consequences of the formation of [Leu5]-enkephalin from the smaller dynorphins are discussed.

Proliferation of immature T cells within the splenocytes of athymic mice by Pseudomonas exotoxin A.

Pseudomonas aeruginosa exotoxin A has been shown to stimulate splenocytes from athymic nude mice. The present studies were performed to characterize the exotoxin A-responsive cells within the splenocytes of athymic nude mice. The results of these studies indicate that exotoxin A-responsive cells were represented in the nylon wool-adherent cell population and in the Ig-depleted splenocyte population (IgNA). Flow microfluorimetry analysis indicated that exotoxin A induced the expansion of Thy1+, CD3-, CD4-, and CD8- cells which also expressed the heat-stable antigen (HSA), an antigen expressed on immature T lymphocytes. Depletion of Thy1+ cells abrogated the exotoxin A-induced proliferation; however, depletion of CD4+ and CD8+ cells did not affect the exotoxin A-induced proliferation of IgNA cells. Thus, the results indicate that exotoxin A stimulates the proliferation of immature T cells within the splenocytes of nude mice that are Thy1+, HSA+, CD3-, CD4-, CD8-.