A voltage-dependent K+ current contributes to membrane potential of acutely isolated canine articular chondrocytes

The electrophysiological properties of acutely isolated canine articular chondrocytes have been characterized using patch-clamp methods. The 'steady-state' current–voltage relationship ( I–V ) of single chondrocytes over the range of potentials from −100 to +40 mV was highly non-linear, showing strong outward rectification positive to the zero-current potential. Currents activated at membrane potentials negative to −50 mV were time independent, and the I–V from −100 to −60 mV was linear, corresponding to an apparent input resistance of 9.3 ± 1.4 GΩ ( n = 23). The outwardly rectifying current was sensitive to the K⁺ channel blocking ion tetraethylammonium (TEA), which had a 50% blocking concentration of 0.66 m m (at +50 mV). The 'TEA-sensitive' component of the outwardly rectifying current had time- and membrane potential-dependent properties, activated near −45 mV and was half-activated at −25 mV. The reversal potential of the 'TEA-sensitive' current with external K⁺ concentration of 5 m m and internal concentration of 145 m m , was −84 mV, indicating that the current was primarily carried by K⁺ ions. The resting membrane potential of isolated chondrocytes (−38.1 ± 1.4 mV; n = 19) was depolarized by 14.8 ± 0.9 mV by 25 m m TEA, which completely blocked the K⁺ current of these cells. These data suggest that this voltage-sensitive K⁺ channel has an important role in regulating the membrane potential of canine articular chondrocytes.     

Distinct hemagglutinin and neuraminidase epitopes involved in antigenic variation of recent human parainfluenza virus type 2 isolates.

A panel of fourteen neutralizing anti-HN monoclonal antibodies (mAbs) to the prototype Greer strain of human parainfluenza virus type 2 (PI2) was used to determine the extent of antigenic variation in recent virus isolates. Competitive binding analysis with the mAbs indicated the presence of at least five distinct antigenic sites (I to V) on the HN glycoprotein molecule. MAbs recognizing different antigenic sites were found to be associated with the hemagglutinin (sites I, IV and V), hemagglutinin and neuraminidase (site II), or neuraminidase (site III) activities. The location of two distinct epitopes identifying the neuraminidase sites (II and III) was further verified from the generation of escape mutants. Antibodies directed to sites I and III failed to show any detectable binding or neutralizing activity against a number of natural PI2 virus isolates collected in Texas between 1986 and 1987. Interestingly, these natural variants, unlike the prototype virus, did not show any detectable neuraminidase activity with fetuin as a substrate and the enzyme activity was only detected with N-acetylneuramin-lactose as an alternative substrate. Despite the observed variation in the antigenic sites, primary infection with the prototype virus or the natural variants generated a protective immune response against challenge infection with the other virus strains.     

Developing health science students into integrated health professionals: a practical tool for learning

Background  

An integrated sense of professionalism enables health professionals to draw on relevant knowledge in context and to apply a set of professional responsibilities and ethical principles in the midst of changing work environments [1, 2]. Inculcating professionalism is therefore a critical goal of health professional education. Two multi-professional courses for first year Health Science students at the University of Cape Town, South Africa aim to lay the foundation for becoming an integrated health professional [3]. In these courses a diagram depicting the domains of the integrated health professional is used to focus the content of small group experiential exercises towards an appreciation of professionalism. The diagram serves as an organising framework for conceptualising an emerging professional identity and for directing learning towards the domains of 'self as professional' [4, 5].     

Mechanism of fluconazole resistance in Candida albicans biofilms: phase-specific role of efflux pumps and membrane sterols

Candida albicans biofilms are formed through three distinct developmental phases and are associated with high fluconazole (FLU) resistance. In the present study, we used a set of isogenic Candida strains lacking one or more of the drug efflux pumps Cdr1p, Cdr2p, and Mdr1p to determine their role in FLU resistance of biofilms. Additionally, variation in sterol profile as a possible mechanism of drug resistance was investigated. Our results indicate that parent and mutant strains formed similar biofilms. However, biofilms formed by double and triple mutants were more susceptible to FLU at 6 h (MIC = 64 and 16 microg/ml, respectively) than the wild-type strain (MIC > 256 microg/ml). At later time points (12 and 48 h), all the strains became resistant to this azole (MIC > or = 256 microg/ml), indicating lack of involvement of efflux pumps in resistance at late stages of biofilm formation. Northern blot analyses revealed that Candida biofilms expressed CDR and MDR1 genes in all the developmental phases, while planktonic cells expressed these genes only at the 12- and 48-h time points. Functionality of efflux pumps was assayed by rhodamine (Rh123) efflux assays, which revealed significant differences in Rh123 retention between biofilm and planktonic cells at the early phase (P = 0.0006) but not at later stages (12 and 48 h). Sterol analyses showed that ergosterol levels were significantly decreased (P < 0.001) at intermediate and mature phases, compared to those in early-phase biofilms. These studies suggest that multicomponent, phase-specific mechanisms are operative in antifungal resistance of fungal biofilms.     

Development and Evaluation of an Interactive CD-ROM Refusal Skills Program to Prevent Youth Substance Use: “Refuse to Use”

An interactive CD-ROM program designed to reduce adolescent substance use was developed and evaluated. The program uses video vignettes to teach refusal skills and socially acceptable responses to substance use situations, specifically offers of marijuana. In a randomized pretest-to-posttest experiment with 74 public school students from six classes in three high schools, significant changes were observed at posttest on (1) the adolescent's personal efficacy to refuse the offer of marijuana, (2) the adolescent's intention to refuse marijuana if offered, and (3) the adolescent's perceptions of the social norms associated with substance use and the importance of respecting another's decision to refuse a drug offer. In addition, adolescents in the treatment condition were able to recall approximately 50% of the portrayed refusal strategies. Findings are discussed with regard to the potential benefits of an interactive multimedia approach for conducting substance use interventions.     

Differential effects of cyclosporine A on Langerhans cells and regulatory T-cell populations in severe psoriasis: an immunohistochemical and flow cytometric analysis

Systemic administration of cyclosporine A (Cy-A; initial dose 5 or 2.5 mg/kg/day) to patients with severe chronic plaque psoriasis produced marked reductions in psoriasis area and severity index within 4 weeks. The clinical response was accompanied, within 1 week, by progressive reductions in T-cell subpopulations (CD3+ and CD4+) and in numbers of interleukin-2 receptor (IL-2-R)-positive (CD25+) cells within lesional skin. Over the first 4 weeks of treatment, these changes were accompanied by reductions in DR+ cells within the epidermis (minor) and dermis (substantial). In contrast, numbers of epidermal CD1+ cells increased substantially during resolution of the skin lesions. Unlike lesional skin, however, no significant changes in absolute numbers of circulating immunoregulatory T-cell populations, including helper/inducer (CD45R) and suppressor/inducer (CD29W) subsets, quantified by dual immunofluorescence labelling, were detected. Moreover, numbers of blood-borne HLA-DR, IL-2-R and transferrin receptor (CD71) positive lymphocytes were unaffected by Cy-A therapy, nor were any differences detected between psoriatic patients and normal controls using these cell markers. Our data suggest that the immunoregulatory effects of Cy-A in psoriasis are mediated via lesional T lymphocytes and that epidermal CD1+ DR- dendritic cells may play an influential role in the regulation of T-cell function and keratinocyte growth during resolution of the skin lesions.     

Reduction of ECS produced retrograde amnesia by post-trial introduction of strychnine

Strychnine sulfate was used to reduce the amount of retrograde amnesia induced by electroconvulsive shock (ECS). Male Swiss-Webster mice were trained on the step-down passive avoidance task, given ECS and then injected intraperitoneally with either 0.1 mg/kg strychnine or saline. The animals were tested in the apparatus one to twenty-four hours later. Mice given ECS alone displayed partial memory for the training (by responding with longer latencies) if tested within two hours after training, but did not display retention twenty-four hours later. Animals given strychnine following ECS had significantly longer step-down latencies at twenty-four hours than did the saline or no injection controls. Further experiments replicated the basic results, and also showed that strychnine injections delayed two hours after the training trial attenuated ECS-induced retrograde amnesia, but that a three hour injection delay was ineffective. The results were interpreted in terms of changes in short-term and long-term memory.     

Interacting effects of temperature and extracellular calcium on the spontaneous release of transmitter at the frog neuromuscular junction

1. Temperature has a characteristic effect on the frequency of m.e.p.p.s at the frog neuromuscular junction; the spontaneous release of transmitter is not affected by temperature changes below 10 degrees C whereas the system is highly temperature-sensitive above 20 degrees C.2. A very similar result is obtained when the experiment is repeated in saline containing Ca(2+) buffered at 5 x 10(-7)M, suggesting that it is unlikely that the major action of temperature is to cause an increase in Ca(2+) influx.3. It is suggested that the main effect of temperature at the presynaptic terminals is a modification of [Ca(2+)](i) by an action on intracellular Ca(2+) stores.4. The interacting effects of theophylline and the divalent cation ionophore A23187 on m.e.p.p. frequency suggest that intracellular Ca(2+) stores, in addition to the mitochondria, may well be of importance in controlling [Ca(2+)](i).5. Changes in [Ca(2+)](o) produce a modification of m.e.p.p. frequency, but the details of the response are dependent on temperature. The spontaneous release of transmitter is most sensitive to an increase in [Ca(2+)](o) at 23 degrees C, whereas the greater effect is found at 13 degrees C when [Ca(2+)](o) is lowered.6. It is suggested (i) that m.e.p.p. frequency is primarily determined by [Ca(2+)](i) at the presynaptic terminals, (ii) that the presynaptic terminals are normally able to maintain [Ca(2+)](i) almost constant in spite of increases in Ca influx associated with ionophore treatment or with a rise in [Ca(2+)](o). However, if the steady-state position of [Ca(2+)](i) is previously raised by an increased efflux from intracellular stores (produced by elevated temperature or theophylline pre-treatment), increased influx causes a rise in both [Ca(2+)](i) and in m.e.p.p. frequency.     

Direct measurement of the thermal responses of nude resting men in dry environments

Summary Two nude resting men were exposed for two-hour periods to each of 25 dry environments, with air temperatures ranging between 12.8° C and 49.1° C and wind speeds between 0.67 m/sec and 4.94 m/sec. The mean radiant temperature of the surroundings was kept equal to the air temperature. Rates of radiant and convective heat exchange were measured directly, separately and continuously. The men had reached a thermal steady state after 105 min in the warm environments, but not in the cold environments. Graphs are presented to show the effect of ambient temperature and wind speed on the radiation and convection rates attained after 105 min, as well as on metabolic rate, sweat evaporation rate, rectal temperature and mean skin temperature. These graphs revealed some important aspects of the behaviour of man's thermal control system. In particular the physiological conductance increased with increasing ambient temperature and then saturated at an ambient temperature near 35° C. This saturation resulted in a constant difference between rectal temperature and mean skin temperature irrespective of the environmental conditions.Published with the permission of the Chamber of Mines of South Africa.     

Immunohistochemistry of neurone specific enolase with gamma subunit specific anti-peptide monoclonal antibodies.

AIMS--To investigate the application in immunohistochemistry of gamma-subunit specific anti-peptide monoclonal antibodies to human neurone specific enolase (NSE); and to determine their reactivity with formalin fixed, wax embedded sections of normal tissue and neuroendocrine tumours. METHODS--Immunohistochemical staining was performed on sections of formalin fixed, wax embedded tissue with two monoclonal antibodies (NSE-P1 and NSE-P2) raised against different synthetic peptides specific for the gamma subunit of human enolase (neurone specific enolase). RESULTS--Both antibodies gave strong immunostaining in normal tissues and cells known to contain NSE. There was no immunoreactivity in tissues containing either the alpha alpha or beta beta isozymes of enolase. The reactivity of the antibodies with a range of neuroendocrine tumours was also studied and both antibodies gave strong immunostaining of tumour cells in the different tumours. CONCLUSIONS--The use of synthetic peptides from defined regions of a molecule as immunogenes provides antibodies of high specificity. These monoclonal antibodies to NSE are ideally suited for immunohistochemical studies and they should be particularly useful in histopathology as they react with epitopes which are resistant to formalin fixation and wax embedding.