N-methyl-D-aspartate triggers neonatal rat hypoglossal motoneurons in vitro to express rhythmic bursting with unusual Mg2+ sensitivity.

The brainstem nucleus hypoglossus innervates the tongue which must contract rhythmically during respiration, chewing and swallowing. Such rhythmic discharges are due to network bursting mediated by AMPA receptor-dependent glutamatergic transmission. The contribution by hypoglossal motoneurons themselves to rhythmicity remains, however, unclear as they might simply express cyclic patterns produced by premotoneurons or, in analogy to spinal motoneurons, might participate to bursting due to activation of their N-methyl-D-aspartate (NMDA) receptors. Using patch clamp recording from hypoglossal motoneurons in slice preparations of neonatal rat brainstem, we observed that NMDA directly depolarized motoneurons to generate various discharge patterns. Most motoneurons produced transient bursts which were consistently restored by repolarizing membrane potential to rest. Fewer motoneurons generated either sustained bursting or random firing. Rhythmic bursts were recorded from XII nerve rootlets even when single motoneuron bursting required hyperpolarization. NMDA evoked bursts were blocked by the Ca2+ antagonist Cd2+, the gap junction blocker carbenoxolone, or Mg2+ free solution, and partially inhibited by tetrodotoxin or nifedipine. Under voltage clamp, NMDA-induced bursting persisted at negative or positive potentials and was resistant to high extracellular Mg2+ in accordance with the observation of widespread motoneuron expression of NMDA 2D receptor subunits that confer poor Mg2+ sensitivity. It is proposed that NMDA depolarized motoneurons with the contribution of Mg2+ insensitive channels, and triggered bursting via cyclic activation/deactivation of voltage-dependent Na+, Ca2+ and K+ currents spread through gap junctions. The NMDA-evoked bursting pattern was similar to the rhythmic discharges previously recorded from the XII nerve during milk sucking by neonatal rats.

Alopecia areata and autoimmunity.

AIM: The aim of the present study was to estimate the association between alopecia areata (AA) and autoimmunity. To such scope, the authors have estimated the presence of autoantibodies in patients with alopecia areata joints to the authors' observation in the last two years. METHODS: In 63 patients affected with different forms of AA (45 cases with multiple patches, 10 with single patch, 5 with alopecia totalis and 3 with alopecia universalis), has been estimated a autoantibodies panel. In 28 subjects (44%) it has been evidenced the presence of autoantibodies; in particular, 23 subjects were affected from AA with multiple patches, 3 from single patch, 1 from alopecia totalis and 1 from alopecia universalis. RESULTS: The autoantibodies against thyroperoxidase and thyroglobulin were found more frequently, followed from antimicrosomal and antinuclear. Except in 5 subjects with autoimmune disease (4 from thyroiditis and 1 from subacute cutaneous lupus erythematosus), all the other cases turned out positives were asymptomatic, then the skin behaved like ''organ proof'' of an organ pathology. On this purpose, in 17 cases (27%) it has been revealed an autoimmune thyroiditis. CONCLUSION: AA is often associated to other autoimmune diseases and that in the 36% of the cases it has permitted to reveal a silent pathology of organ.

The effects induced by the sulphonylurea glibenclamide on the neonatal rat spinal cord indicate a novel mechanism to control neuronal excitability and inhibitory neurotransmission.

BACKGROUND AND PURPOSE: Using the neonatal rat spinal cord in vitro, we investigated the action of glibenclamide, a drug possessing dual pharmacological effects, namely block of K(ATP) channels and of the cystic fibrosis transmembrane conductance regulator (CFTR). EXPERIMENTAL APPROACH: Intra- and extracellular recordings were performed on motoneurons and interneurons. RT-PCR and western immunoblotting were used to determine gene and protein expression. KEY RESULTS: Glibenclamide (50 microM) facilitated mono- and polysynaptic reflexes, hyperpolarized motoneuron resting potential, increased action potential amplitude, decreased Renshaw cell-mediated recurrent inhibition, and increased network excitability by depressing GABA- and glycine-mediated transmission. The action of glibenclamide was mimicked by tolbutamide (500 microM) or the CFTR blocker diphenylamine-2,2-dicarboxylic acid (500 microM). The action of glibenclamide was independent from pharmacological inhibition of the Na(+)-K(+) pump with strophanthidin (4 microM) and was associated with a negative shift in the extrapolated reversal potential for CI(-) dependent synaptic inhibition. On interneurons, intracellularly-applied 8-bromo-cAMP elicited an inward current and resistance decrease; effects antagonized by the selective CFTR antagonist, CFTR(inh)-172 (5 microM). RT-PCR and western immunoblotting indicated strong expression of the CFTR in neonatal rat spinal cord. CONCLUSIONS AND IMPLICATIONS: These data suggest the CFTR expressed in motoneurons and interneurons of the neonatal spinal cord is involved in the control of Cl(-) homeostasis and neuronal excitability. CFTR appeared to contribute to the relatively depolarized equilibrium potential for synaptic inhibition, an important process to control hyperexcitability and seizure-predisposition in neonates.

Early signs of motoneuron vulnerability in a disease model system: Characterization of transverse slice cultures of spinal cord isolated from embryonic ALS mice.

Mutations in the SOD1 gene are associated with familial amyotrophic lateral sclerosis. The mechanisms by which these mutations lead to cell loss within the spinal cord ventral horns are unknown. In the present report we used the G93A transgenic mouse model of amyotrophic lateral sclerosis to develop and characterize an in vitro tool for the investigation of subtle alterations of spinal tissue prior to frank neuronal degeneration. To this aim, we developed organotypic slice cultures from wild type and G93A embryonic spinal cords. We combined immunocytochemistry and electron microscopy techniques to compare wild type and G93A spinal cord tissues after 14 days of growth under standard in vitro conditions. By SMI32 and choline acetyl transferase immunostaining, the distribution and morphology of motoneurons were compared in the two culture groups. Wild type and mutant cultures displayed no differences in the analyzed parameters as well as in the number of motoneurons. Similar results were observed when glial fibrillary acidic protein and myelin basic protein-positive cells were examined. Cell types within the G93A slice underwent maturation and slices could be maintained in culture for at least 3 weeks when prepared from embryos. Electron microscopy investigation confirmed the absence of early signs of mitochondria vacuolization or protein aggregate formation in G93A ventral horns. However, a significantly different ratio between inhibitory and excitatory synapses was present in G93A cultures, when compared with wild type ones, suggesting the expression of subtle synaptic dysfunction in G93A cultured tissue. When compared with controls, G93A motoneurons exhibited increased vulnerability to AMPA glutamate receptor-mediated excitotoxic stress prior to clear disease appearance. This in vitro disease model may thus represent a valuable tool to test early mechanisms contributing to motoneuron degeneration and potential therapeutic molecular interventions.

[A vaccination campaign for gypsy children in Rome]. / Una campagna vaccinale in favore dei bambini Rom e Sinti a Roma.

Several studies conducted in Rome have shown low levels of vaccination coverage in gypsy communities. Thus a vaccination campaign targeting to 2400 gypsy children aged 0-13 years, present in 32 settlements in Rome, was conducted in 2002. The campaign was related to vaccinations required and recommended in Italy: diphtheria, tetanus, hepatitis B, pertussis (whooping cough), haemophilus influenzae and measles. In the majority of cases it was decided to carry out the vaccinations directly in the gypsy settlements, in accordance with the methods of pulse immunisation. In the case of small settlements a strategy for reorienting the population to vaccination centres was adopted. Around 2000 children were vaccinated, equivalent to 80% of the paediatric population present during the period. The number of children who have never been vaccinated has decreased from 40% prior to the campaign to 9% after the third week. Vaccination coverage in medium- and small-sized settlements (<200 inhabitants) after the campaign shows values of over 70%; in the large settlements, more modest increases have been recorded and coverage has rarely exceeded 50%. This experience has highlighted the importance of networking between public healthcare institutions and non-profit organisations. The mobilisation of a wide range of competences has thus enabled the attainment of a high level of effectiveness.

Mechanism of mechanically induced intercellular calcium waves in rabbit articular chondrocytes and in HIG-82 synovial cells.

Intercellular communication through gap junctions allows tissue coordination of cell metabolism and sensitivity to extracellular stimuli. Intercellular Ca2+ signaling was investigated with digital fluorescence video imaging in primary cultures of articular chondrocytes and in HIG-82 synovial cells. In both cell types, mechanical stimulation of a single cell induced a wave of increased Ca2+ that was communicated to surrounding cells. Intercellular Ca2+ spreading was inhibited by 18alpha-glycyrrhetinic acid, demonstrating the involvement of gap junctions in signal propagation. In the absence of extracellular Ca2+, mechanical stimulation induced communicated Ca2+ waves similar to controls; however, the number of HIG-82 cells recruited decreased significantly. Mechanical stress induced Ca2+ influx both in the stimulated chondrocyte and HIG-82 cell, but not in the adjacent cells, as assessed by the Mn2+ quenching technique. Treatment of cells with thapsigargin and with the phospholipase C (PLC) inhibitor U73122 blocked mechanically induced signal propagation. These results provide evidence that in chondrocytes and in HIG-82 synovial cells, mechanical stimulation activates PLC, thus leading to an increase of intracellular inositol 1,4,5-trisphosphate. The second messenger, by permeating gap junctions, stimulates intracellular Ca2+ release in neighboring cells. It is concluded that intercellular Ca2+ waves may provide a mechanism to coordinate tissue responses in joint physiology.

A population-based study on the frequency of additional congenital malformations in infants with congenital hypothyroidism: data from the Italian Registry for Congenital Hypothyroidism (1991-1998).

In the last decade a high frequency of other congenital anomalies has been reported in infants with congenital hypothyroidism (CH) detected by neonatal screening. In the present study the occurrence of additional congenital malformations (CM) in the population of CH infants detected in Italy between 1991 and 1998 (n = 1420) was investigated. In Italy all of the centers in charge of screening, treatment, and follow-up of CH adhere to the Italian National Registry of infants with CH. In this study a high prevalence of additional CM (8.4%), more than 4-fold higher than that reported in the Italian population (1-2%), was found in the population of CH infants. Cardiac anomalies represented the most frequent malformations associated with CH, with a prevalence of 5.5%. However, a significant association between CH and anomalies of nervous system, eyes, and multiple CM was also observed. In conclusion, the significantly higher frequency of extrathyroidal congenital malformations reported in the CH infants than in the general population represents a further argument supporting the role of a genetic component in the etiology of CH. Investigations of the molecular mechanisms underlying developmental events of formation of thyroid and other organs represent critical steps in the knowledge of CH etiology.

Epidemiology of legal abortion in Italy.

The paper reports on the trends of induced abortion in Italy since its legalization in 1978. Data collected through the national surveillance system show that the abortion rates and ratios in Italy, after an initial increase, have been stable in the last four years, and are comparable to those of other European countries. Italian women seeking abortion, however, are generally older and more frequently married and parous than in most other western countries. The data on abortion services and procedures show a slow improvement with increasing use of local rather than general anaesthesia, suction rather than surgical curettage, and outpatient rather than inpatient admissions. Post-abortion complications are infrequent and there have been no cases of death from legal abortion. The paper attempts to analyse the effect of legal abortion on fertility and on other maternal and child health indices, reaching tentative conclusions on the contribution of the legalization of abortion to the improvement of reproductive health. Uneven availability of abortion services, however, is currently a major problem; in many of the less developed areas of the country, it is estimated that as many as 50% of abortions are still performed outside the legal abortion service network.

Measles epidemiology in Italy.

In preparation for a mass vaccination programme, the immune status with regard to measles was determined in over 8300 unvaccinated children aged 0-13 years, residing in eight Italian cities with different socioeconomic situations and geographical locations. The age corresponding to the 50% prevalence of immunes appeared to be intermediate (2.9-5.5 years) between that reported for industrialized (6-7 years) and developing countries (1-2 years). The 50% prevalence of natural immunity was reached at an earlier age in southern cities in which poorer socioeconomic and hygienic conditions prevailed; the earlier occurrence of measles in these areas was confirmed by a more detailed serological study of children in the first 24 months of life. For children aged 2-13 years, serological results showed that the history of measles reported by parents on questionnaires gave high positive predictive values (over 85%). Our seroepidemiological study shows that, on the basis of the ages of 25 and 75% prevalence of immunes, the target population for a mass immunization programme in Italy can be assumed to be aged from 12 months to 7 years. However, special attention should be given to the poorest areas, especially in southern Italy, where measles occurs earlier and can be particularly severe.

Non-linear dependence of the dielectric properties of chick embryo myoblast membranes exposed to a sinusoidal 50 Hz magnetic field.

Primary chick embryo myoblasts can be a useful tool for studying the developmental events which accompany myoblast differentiation, particularly myoblast membrane fusion. To determine whether the electrical properties and/or fusion in these systems are affected by 50 Hz magnetic fields, chick embryo myoblast cultures were exposed to B-field intensities ranging from 1 to 10 mT. The electrical parameters of the myoblasts, i.e. membrane conductivity, membrane permittivity and the conductivity of the cell interior (cytosol) were determined by the analysis of conductivity dispersion data in the radio frequency range (10 kHz-100 MHz). Preliminary results indicate that the time of fusion (60 h) is not affected by these fields, but that the absolute values of the two membrane electrical parameters are affected. In particular, a B-field intensity-dependent decrease was observed. The maximum effect resulted after a 1 h exposure to a magnetic flux density of about 5 mT. The conductivity of the cytosol remained unchanged. These data seem to indicate that exposure to 50 Hz magnetic fields affects both static and dynamic membrane properties in primary chick embryo myoblasts.

A static magnetic field does not affect the dielectric properties of chick embryo myoblast membranes.

We have recently demonstrated using dielectic relaxation studies in the radiowave frequency range that sinusoidal 50 Hz magnetic fields (with intensities ranging from 1 to 10 mT) induce a nonlinear change in both membrane conductivity and permittivity of primary chick embryo myoblasts in vitro. It was the aim of the present study to determine if a DC-induced static magnetic field is capable of generating similar variations in the membrane conductivity and/or the membrane permittivity of chick embryo myoblasts. The results indicate that when the myogenic cells are exposed to a static magnetic field of either 1, 3 or 5 mT (values comparable with the previous extremely low frequency study), no changes in the membrane electrical parameters can be observed with respect to controls. Differences in the characteristics of static and extremely low frequency fields as well as the possible mechanisms underlying the contrasting results with these two types of magnetic fields are discussed.

Human natural killer cytotoxic activity is not affected by in vitro exposure to 50-Hz sinusoidal magnetic fields.

Epidemiological studies have suggested, but not demonstrated, a role of exposure to 50/60-Hz magnetic fields in increasing cancer risk in man (workers and the general population). A possible target of magnetic fields is the immune system. In particular, it is known that an important defence against cancer is represented by natural killer (NK) cells capable of killing cancer cell targets. To test this hypothesis, human NK cells, stimulated or not with phytohaemagglutinin or interleukin 2, were exposed to 50-Hz sinusoidal magnetic fields before or during the cytotoxicity test, and then mixed with a variety of target cancer cell lines (Daudi, Raji, U937, H14, IGROV, SW626, K562, HL60). The experiments were performed in two laboratories (Rome and Modena) by means of two different exposure systems. The results of both laboratories suggest that 50-Hz sinusoidal magnetic fields with flux densities up to 10 mT do not affect the cytotoxic activity of human NK cells.

Sex education in secondary schools: an Italian experience.

PURPOSE: To evaluate the impact on Italian teenagers' knowledge and attitudes of an experimental program in sex education. METHODS: The program consisted of five workshops in five public secondary schools in Rome. The impact of the program was evaluated with three questionnaires designed for this study. A pretest evaluated baseline knowledge, a posttest measured the gain in knowledge, and a third questionnaire estimated retention of information 4-6 months later. Three hundred seventy-six students attending secondary school participated in the program. Thirteen questions were selected for the analysis, and improvement was measured by the difference between pretest and posttest scores. RESULTS: Ninety-six percent of the sample was in favor of sex education programs in schools. The students had an average 50% gain in the percentage of correct answers following the course. The mean score based on the number of correct answers to questions showed a baseline value of 5.00, a posttest value of 11.80, and a test value 4-6 months later of 10.7. Results showed students' positive attitude toward school sex education courses, low baseline knowledge, and a good ability to learn. CONCLUSIONS: In Italy there is a need for collaboration between schools and local health services to promote knowledge and prevention in reproductive health among teenagers.

Reducing pain of first trimester abortion under local anaesthesia.

The study aimed to estimate the pain related to first trimester abortion under local and general anaesthesia. One hundred and eighty-one women undergoing abortion of first trimester under local anaesthesia and 198 under general anaesthesia were interviewed 1 h after the end of the procedure. About 50% of all women described their experience as less painful than expected. Over 50% of the women described the pain during the procedure as mild or moderate according to the verbal rating scale. The step-up logistic regression analyses identified four important variables: time interval less than 2 min from local anaesthesia injection to the beginning of the procedure (RR = 3), lack of choice between local and general anaesthesia (RR = 1.9), history of frequent use of analgesics (RR = 1.9) and nulliparity (RR = 1.7). These data indicate that skillfully performed abortion with local anaesthesia is a procedure tolerated by most women. Moreover, because it carries lower risk of complications and costs less, its use should be encouraged.

PIP: The World Health Organization has urged a reduction in the number of first-trimester induced abortions performed under general anesthesia. To determine the implications of this recommendation for pain control, 181 women undergoing first-trimester abortion under local anesthesia and 198 aborted under general anesthesia at University Hospital in Rome, Italy, were interviewed 1 hour after the procedure. Pain intensity was assessed through both the visual analogue and verbal rating scales. Women who selected or were assigned to general anesthesia tended to be younger (mean age, 27 years) and nulliparous. Over 50% of local anesthesia patients described their pain during the procedure as mild or moderate, 30% rated it severe, and 18% characterized it as distressing but time-limited. 60 minutes after the abortion, pain scores were higher in the general anesthesia group. Logistic regression analysis identified four significant predictors of pain: time interval less than 2 minutes from local anesthetic injection to the beginning of the procedure (relative risk (RR), 3.0), lack of choice between general or local anesthesia (RR, 1.9), history of frequent use of analgesics (RR, 1.9), and nulliparity (RR, 1.7). 91% of general anesthesia and 73% of local anesthesia patients stated they would recommend their method of anesthesia to a friend undergoing abortion. These findings suggest that local anesthesia, less costly and with a lower risk of complications than general anesthesia, is feasible for pain control, especially if administered 2-5 minutes before the procedure begins.

Do antenatal classes benefit the mother and her baby?

OBJECTIVE: To examine the characteristics of women attending antenatal classes and evaluate the effects of these classes on mothers' and babies' health. METHODS: A population-based observational study on care during pregnancy, delivery and in the postnatal period was carried out in 1995-96. A total of 9004 women resident in 13 regions of Italy who delivered in a 4-month period were interviewed. The outcomes studied were attendance at antenatal classes, Cesarean section, bottle feeding, satisfaction with the experience of childbirth, knowledge of contraception, breast feeding and baby care. RESULTS: A total of 2065 (23.0%) women attended antenatal classes. Women without previous children, those with a higher level of education and office workers were more likely to attend classes. Women who attended antenatal classes had a much lower risk of Cesarean section and were about half as likely to bottle feed while in hospital compared with non-attenders. They received better information on contraception, breast feeding and baby care. Women who attended classes and applied the techniques learned were more satisfied with the experience of childbirth. CONCLUSION: Antenatal classes seem to improve women's knowledge and competence. This may provide a defence against the tendency to overmedicalize pregnancy and childbirth.

Intercellular Ca2+ waves in mechanically stimulated articular chondrocytes.

Articular cartilage is a tissue designed to withstand compression during joint movement and, in vivo, is subjected to a wide range of mechanical loading forces. Mechanosensitivity has been demonstrated to influence chondrocyte metabolism and cartilage homeostasis, but the mechanisms underlying mechanotransduction in these cells are poorly understood. In many cell types mechanical stimulation induces increases of the cytosolic Ca2+ concentration that propagates from cell to cell as an intercellular Ca2+ wave. Cell-to-cell communication through gap junctions underlies tissue co-ordination of metabolism and sensitivity to extracellular stimuli: gap junctional permeability to intracellular second messengers allows signal transduction pathways to be shared among several cells, ultimately resulting in co-ordinated tissue responses. Mechanically-induced Ca2+ signalling was investigated with digital fluorescence video imaging in primary cultures of rabbit articular chondrocytes. Mechanical stimulation of a single cell, obtained by briefly distorting the plasmamembrane with a micropipette, induced a wave of increased Ca2+ that was communicated to surrounding cells. Intercellular Ca2+ spreading was inhibited by 18 alpha-glycyrrhetinic acid, suggesting the involvement of gap junctions in signal propagation. The functional expression of gap junctions was assessed, in confluent chondrocyte cultures, by the intercellular transfer of Lucifer yellow dye in microinjection experiments while the expression of connexin 43 could be detected in Western blots. A series of pharmacological tools known to interfere with the cell calcium handling capacity were employed to investigate the mechanism of mechanically-induced Ca2+ signalling. In the absence of extracellular Ca2+ mechanical stimulation induced communicated Ca2+ waves similar to controls. Mechanical stress induced Ca2+ influx both in the stimulated chondrocyte but not in the adjacent cells, as assessed by the Mn2+ quenching technique. Cells treatment with thapsigargin and with the phospholipase C inhibitor U73122 blocked mechanically-induced signal propagation. These results provide evidence that in chondrocytes mechanical stimulation activates phospholipase C, thus leading to an increase of intracellular inositol 1,4,5-trisphosphate. The second messenger, by permeating gap junctions, stimulates intracellular Ca2+ release in neighbouring cells. Intercellular Ca2+ waves may provide a mechanism to co-ordinate tissue responses in cartilage physiology.

[International guidelines on the diagnostic applications of magnetic resonance. International Non-Ionizing Radiation Committee of the International Radiation Protection Association]. / Linee guida internazionali nelle applicazioni diagnostiche della risonanza magnetica. Comitato Internazionale per le Radiazioni Non Ionizzanti dell'Associazione Internazionale per le Protezioni Radiologiche.

Magnetic resonance imaging has become an established diagnostic modality. The clinical usefulness of in vivo magnetic resonance spectroscopy was demonstrated in several instances and is being explored further. These techniques involve exposure of the patient to static and time-varying magnetic fields and radiofrequency electromagnetic fields. In particular exposure situations, these fields may pose a health hazard. The purpose of this paper is to provide information on the guidelines issued by the International Non-Ionizing Radiation Committee of the International Radiation Protection Association on levels of exposure and health effects from magnetic and radiofrequency electromagnetic fields associated with magnetic resonance diagnostic devices, and on precautions to be taken to minimize health hazards to patients undergoing magnetic resonance examinations.

[Ethical aspects of strategies for prevention]. / Gli aspetti etici nella strategia per la prevenzione.

Prevention must be offered actively to all sections of the target population. The spontaneous process select people at lower risk. The active offer of preventive measures must be modulated on the difficulty to be reached of the different sections of the target population. The main problem of prevention is that the risk factors of non compliance are the same for a higher incidence or prevalence of events or conditions. The efficacy of a preventive strategy is valuable observing a reduction of the incidence or prevalence of preventable events or conditions in a definite period of time and in a defined community. In the case of prevention ethical and professional quality coincide.

[Family health services and preventive strategies]. / I consultori familiari e le strategie di prevenzione.

Health promotion is effective if it is based on the active offer of preventive measures to the target population. The active offer must be sustained by non standardised communication procedures, with the aim of producing empowerment of involved people. People, who are socially deprived and therefore at high risk, are hard to reach. They need special attention and resources. Prevention services and, in particular, those devoted to women, children and adolescents must be sufficiently diffused among the population. They must change from being dependent on demand to being based on active offer of preventive measures.

[Evaluation of the impact of birth preparation courses on the health of the mother and the newborn]. / Valutazione degli effetti dei corsi di preparazione alla nascita sulla salute della madre e del neonato.

In 1995-96 a knowledge, attitude and practise (KAP) survey on care during pregnancy, delivery and in the post-natal period was carried out by the National Institute of Health in Italy. Of the 9004 women interviewed, 23% have attended antenatal classes (36% in hospitals, 51% in maternal and child health, MCH, clinics and 13% privately). Women attending these classes tend to be well-educated, primigravidae and resident in the North. Women who attend antenatal classes, especially at MCH clinics, have a lower risk of caesarean section (OR = 0.60), of bottle feeding while in hospital (OR = 0.45) and of receiving no information on contraception (OR = 0.31). Women who attend classes and are able to apply the technique learnt have a reduced risk of being dissatisfied with the experience of childbirth (OR = 0.72), whereas the others have an increased risk of dissatisfaction (OR = 1.87).