Alcohol and bone: review of dose effects and mechanisms.

Alcohol is widely consumed across the world. It is consumed in both social and cultural settings. Until recently, two types of alcohol consumption were recognized: heavy chronic alcohol consumption or light consumption. Today, there is a new pattern of consumption among teenagers and young adults namely: binge drinking. Heavy alcohol consumption is detrimental to many organs and tissues, including bones, and is known to induce secondary osteoporosis. Some studies, however, have reported benefits from light alcohol consumption on bone parameters. To date, little is known regarding the effects of binge drinking on bone health. Here, we review the effects of three different means of alcohol consumption: light, heavy, and binge drinking. We also review the detailed literature on the different mechanisms by which alcohol intake may decrease bone mass and strength. The effects of alcohol on bone are thought to be both direct and indirect. The decrease in bone mass and strength following alcohol consumption is mainly due to a bone remodeling imbalance, with a predominant decrease in bone formation. Recent studies, however, have reported new mechanisms by which alcohol may act on bone remodeling, including osteocyte apoptosis, oxidative stress, and Wnt signalling pathway modulation. The roles of reduced total fat mass, increased lipid content in bone marrow, and a hypoleptinemia are also discussed.

Centrosome fine ultrastructure of the osteocyte mechanosensitive primary cilium.

The centrosome is the principal microtubule organization center in cells, giving rise to microtubule-based organelles (e.g., cilia, flagella). The aim was to study the osteocyte centrosome morphology at an ultrastructural level in relation to its mechanosensitive function. Osteocyte centrosomes and cilia in tibial cortical bone were explored by acetylated alpha-tubulin (AαTub) immunostaining under confocal microscopy. For the first time, fine ultrastructure and spatial orientation of the osteocyte centrosome were explored by transmission electron microscopy on serial ultrathin sections. AαTub-positive staining was observed in 94% of the osteocytes examined (222/236). The mother centriole formed a short primary cilium and was longer than the daughter centriole due to an intermediate zone between centriole and cilium. The proximal end of the mother centriole was connected with the surface of daughter centriole by striated rootlets. The mother centriole exhibited distal appendages that interacted with the cell membrane and formed a particular structure called "cilium membrane prolongation." The primary cilium was mainly oriented perpendicular to the long axis of bone. Mother and daughter centrioles change their original mutual orientation during the osteocyte differentiation process. The short primary cilium is hypothesized as a novel type of fluid-sensing organelle in osteocytes.

A modular spring-loaded actuator for mechanical activation of membrane proteins.

How cells respond to mechanical forces by converting them into biological signals underlie crucial cellular processes. Our understanding of mechanotransduction has been hindered by technical barriers, including limitations in our ability to effectively apply low range piconewton forces to specific mechanoreceptors on cell membranes without laborious and repetitive trials. To overcome these challenges we introduce the Nano-winch, a robust, easily assembled, programmable DNA origami-based molecular actuator. The Nano-winch is designed to manipulate multiple mechanoreceptors in parallel by exerting fine-tuned, low- piconewton forces in autonomous and remotely activated modes via adjustable single- and double-stranded DNA linkages, respectively. Nano-winches in autonomous mode can land and operate on the cell surface. Targeting the device to integrin stimulated detectable downstream phosphorylation of focal adhesion kinase, an indication that Nano-winches can be applied to study cellular mechanical processes. Remote activation mode allowed finer extension control and greater force exertion. We united remotely activated Nano-winches with single-channel bilayer experiments to directly observe the opening of a channel by mechanical force in the force responsive gated channel protein, BtuB. This customizable origami provides an instrument-free approach that can be applied to control and explore a diversity of mechanotransduction circuits on living cells.

G-protein-coupled receptor oligomers: two or more for what? Lessons from mGlu and GABAB receptors.

G-protein-coupled receptors (GPCRs) are key players in the precise tuning of intercellullar communication. In the brain, both major neurotransmitters, glutamate and GABA, act on specific GPCRs [the metabotropic glutamate (mGlu) and GABA(B) receptors] to modulate synaptic transmission. These receptors are encoded by the largest gene family, and have been found to associate into both homo- and hetero-oligomers, which increases the complexity of this cell communication system. Here we show that dimerization is required for mGlu and GABA(B) receptors to function, since the activation process requires a relative movement between the subunits to occur. We will also show that, in contrast to the mGlu receptors, which form strict dimers, the GABA(B) receptors assemble into larger complexes, both in transfected cells and in the brain, resulting in a decreased G-protein coupling efficacy. We propose that GABA(B) receptor oligomerization offers a way to increase the possibility of modulating receptor signalling and activity, allowing the same receptor protein to have specific properties in neurons at different locations.

In-vitro and in-vivo design and validation of an injectable polysaccharide-hydroxyapatite composite material for sinus floor augmentation.

OBJECTIVE: Polysaccharide-based composite matrices consisting of natural polysaccharides, pullulan and dextran supplemented with hydroxyapatite (Matrix-HA) have recently been developed. The principal objective of this study was to evaluate the capacities of this composite material to promote new bone formation in a sinus lift model in the sheep. Secondary objectives were to evaluate in vitro properties of the material regarding cell adhesion and proliferation. METHODS: In this report, once such composite matrix was prepared as injectable beads after dispersion in a physiological buffer, and evaluated using a large animal model (sheep) for a sinus lift procedure. RESULTS: In vitro studies revealed that these microbeads (250-550µm in diameter) allow vascular cell adhesion and proliferation of Endothelial Cells (EC) after 1 and 7 days of culture. In vivo studies were performed in 12 adult sheep, and newly formed tissue was analyzed by Cone Beam Computed Tomography (CBCT scanning electron microscopy (SEM) and by histology 3 and 6 months post-implantation. CBCT analyses at the implantation time revealed the radiolucent properties of these matrices. Quantitative analysis showed an increase of a dense mineralized tissue in the Matrix-HA group up to 3 months of implantation. The mineralized volume over total volume after 6 months reached comparable values to those obtained for Bio-Oss® used as positive control. Histological examination confirmed that the Matrix-HA did not induce any long term inflammatory events, and promoted direct contact between the osteoid tissue and lamellar bone structures and beads. After 6 months, we observed a dense network of osteocytes surrounding both biomaterials as well as a newly vascularized formed tissue in close contact to the biomaterials. SIGNIFICANCE: In conclusion, the absence of animal components in Matrix-HA, the osteoconductive property of Matrix-HA in sheep, resulting in a dense bone and vascularized tissue, and the initial radiolucent property to follow graft integration offer great promises of this composite material for clinical use.

Further characterization of the changes occurring in the plasma lipoprotein spectrum in the European badger (Meles meles L.) during winter.

The plasma lipoprotein pattern in the European badger has been shown previously to undergo marked and complex quantitative and qualitative seasonal modifications (Laplaud, P.M. et al., 1980, J. Lipid Res., 21, 724-738). However, the conventional ultracentrifugal techniques then in use in our laboratory were of insufficient discriminating power with regard to the numerous lipoprotein fractions whose presence was suggested by our analyses. In the present study, a new density gradient ultracentrifugation procedure was applied to the more detailed determination of the distribution of plasma lipoproteins. The first series of analyses was performed in early December and the second in March, i.e. at the dates when the maximum and minimum, respectively, of lipidemia occur in this species. The fractions thus obtained, each of which corresponded to a narrow density interval, were analyzed subsequently for chemical composition, appearance upon polyacrylamide gel electrophoresis, and for their content of tetramethylurea-soluble apolipoproteins in alkaline-urea gels. Changes occurring from December to March included a large decrease in the plasma concentration of the 1.015-1.065 g/ml lipoproteins, chemical analysis of this material being compatible with the presence of at least two lipoprotein populations. On the other hand, high-density lipoproteins (1.065-1.162 g/ml) appeared less variable in chemical composition, although the proportion of those with lower density decreased considerably in early spring. Polyacrylamide gel electrophoresis of the native fractions showed multiple bands in most of them; the tetramethylurea-soluble apoprotein profile remained similar at the two dates considered with an apolipoprotein A-I-like component present in large amounts throughout the entire low- and high-density ranges.

Different responses of plasma ACTH and corticosterone and of plasma interleukin-1 beta to single and recurrent endotoxin challenges.

In a parallel study in 10 individual rats, three time series of plasma concentrations of ACTH, corticosterone (CORT), and interleukin-1 beta (IL-1 beta) were measured before (time 0) and at intervals between 15 and 480 min following intra-arterial (i.a.) infusions of 25 microgram/kg lipopolysaccharide (LPS). All LPS injections were given at 9 AM. The first time series was performed on naive rats (day 1). A sequence of six daily injections (days 3-8) of the same dose of LPS followed. The post-LPS time course of the plasma ACTH, CORT and IL-1 beta levels were studies on days 3 (second injection) and 8 (seventh injection). The first LPS injection induced a rapid (30 min) eightfold rise in plasma ACTH and CORT, culminating in concentrations 30 times the baseline at 60 min (ACTH) and 15 times baseline at 120 min (CORT). Both hormones receded back to the initial basal level at 480 min. On the other hand, IL-1 beta increased slowly to peak at 13 times baseline 120 min before declining to minimal seven- to ninefold basal levels, 480 min and even 48 h post-LPS. During the second phase of the experiment starting 48 h after the initial LPS priming sequence, the ACTH and CORT responses to daily recurrent LPS injections again differed from those of IL-1 beta. The post-LPS time courses of the ACTH and CORT reaction displayed a typical pattern of a progressive attenuation studied at days 3 and 8. The peak amplitudes at days 3 and 8 were reduced to 60 and 10%, respectively, for ACTH, and to 85 and 45% for CORT of those observed at the first LPS test. The duration of the response (both) was also shortened from 480 min (first LPS test) to 300 min at days 3 and 8. The post-LPS patterns of the IL-1 beta responses were characterized, first by basal levels seven to nine times higher than the initial baseline values (day 1), and by a rapid suppression of the post-LPS response, with only a slight (30%) increase at day 3 and no increase at day 8. Thus, after both acute and recurrent LPS administration, ACTH/CORT and IL-1 beta reacted differently to the endotoxin challenge. The two LPS reactive systems were not correlated. This is inconsistent with the often proposed role of increased plasma IL-1 beta release as an intermediary factor in the LPS-induced recruitment of the corticotropic axis in general infections.

A year-long study of changes induced by thyroidectomy in the plasma lipid and lipoprotein spectrum in the European badger.

Hypothyroidism is associated with hypercholesterolemia and increased risk for atherosclerotic disease. The European badger exhibits large seasonal changes in thyroid activity and the annual minimum of plasma thyroxine level in this species occurs at the same period of the year (i.e. late fall) as a pronounced hypercholesterolemia. We examined the plasma lipid and lipoprotein spectrum in a group of thyroidectomized male badgers every month for a year. Non-operated animals were used as controls. Our analyses included measurement of plasma lipid levels, density gradient ultracentrifugation of lipoproteins, electrophoresis of lipoproteins and apolipoproteins, and histological studies. Maximal differences between the two groups of animals were observed during spring, occurring concomitantly with the annual maximum of plasma thyroxine concentration in control badgers. Comparison with the latter animals revealed a permanent hypercholesterolemia and hyperphospholipidemia in thyroidectomized badgers, while their lipoprotein spectrum was characterized by the continual presence of elevated concentrations of cholesterol-rich lipoproteins of d congruent to 1.015 - 1.027 g/ml. The ratio of triglyceride/cholesteryl ester content in such lipoproteins remained constant throughout the year, resembling that noted in intact animals during late fall. Other features distinguishing the lipoprotein spectrum in thyroidectomized badgers were: (1) higher levels of lipoproteins with d 1.027 - 1.065 g/ml and d 1.065 - 1.100 g/ml, and (2) a cholesteryl ester enrichment of both these lipoprotein subclasses. The two groups of animals shared a heterogeneity of low density lipoprotein subfractions isolated on density gradients, together with the presence of apolipoproteins with molecular weights respectively typical of human apolipoproteins A-I and B throughout the low density range. Arterial walls and heart tissues from intact and thyroidectomized animals were free of atherosclerotic lesions at the end of the experimental period.

A year-long study of changes induced by castration in the plasma lipid and lipoprotein spectrum in the European badger.

In man, an influence of male sex hormones on plasma lipid transport is well established; however, recent data on this subject in the literature are both relatively lacking and occasionally conflicting. The male European badger exhibits seasonal variations of large amplitude in its gonadic function. We have therefore attempted to establish the influence of male sex steroids on plasma lipids and lipoproteins in this species. For this purpose, we have examined the plasma lipid and lipoprotein spectrum in a group of castrated male badgers every month for a year, non-operated animals being used as controls. Our analyses included measurement of plasma lipid levels, density gradient ultracentrifugation of lipoproteins, electrophoresis of lipoproteins and apolipoproteins, and evaluation of plasma testosterone and thyroxine levels. The differences observed between the 2 groups of animals were maximal during the months when plasma testosterone was elevated in intact badgers (January to July). For this period, castration resulted in higher plasma concentrations of cholesterol, phospholipids and triglycerides, while the latter alone remained significantly more elevated in operated animals until the end of our experiments. With regard to lipoproteins, the main effect of castration consisted of a large augmentation in the concentration of lipoproteins with d approximately equal to 1.027-1.065 g/ml which were responsible for the transport of most of the increased amounts of triglycerides present in the plasma of castrated badgers. The proportion of apoprotein B in the protein moiety of these lipoprotein components was enhanced after castration. Other changes in the lipoprotein spectrum included (1) a moderate increase in the concentration of lipoproteins with d less than 1.015 g/ml and 1.019-1.027 g/ml, and (2) a modification of the respective proportions of high density lipoproteins with d 1.065-1.100 g/ml and d 1.100-1.162 g/ml. Finally, no considerable differences between the 2 groups of animals were noted in the respective percentages of the various chemical constituents in each lipoprotein subfraction assayed, except for those with d 1.023-1.027 g/ml, which, in castrated badgers, did not exhibit the enrichment in triglycerides usually noted during late winter and spring in intact animals.

Mollicutes DNA polymerases: characterization of a single enzyme from Mycoplasma mycoides and Ureaplasma urealyticum and of three enzymes from Acholeplasma laidlawii.

The DNA polymerase activity of different members of Mollicutes was studied. A single DNA polymerase was found in Mycoplasma mycoides and Ureaplasma urealyticum, type species of the genera Mycoplasma and Ureaplasma, and was compared with the previously described Mycoplasma orale enzyme. Most of their properties were comparable; an immunological relationship was demonstrated between M. orale and M. mycoides enzymes by immunoblotting. In contrast to these results, three different DNA polymerases were purified in Acholeplasma laidlawii, type species of the genus Acholeplasma which, in this aspect, resembles the genus Spiroplasma. A 3'-5' exonuclease activity was found in the different purified preparations. In M. mycoides, M. orale and one of the three A. laidlawii preparations, the 3'-5' exonuclease could be separated from the DNA polymerase by non-denaturing PAGE. The presence of a single DNA polymerase seems to be a typical feature of the Mycoplasmataceae, which include the genera Mycoplasma and Ureaplasma, in contrast to the occurrence of three enzymes within the Acholeplasmataceae and Spiroplasmataceae. These results are in agreement with the phylogenetic tree of Mollicutes proposed from their 5 S and 16 S rRNA sequence comparisons, in which the evolution of Acholeplasma and Spiroplasma branches led, by genome reductions, to Mycoplasma and Ureaplasma species.

Ependymal and choroidal cells in culture: characterization and functional differentiation.

During the past 10 years, our teams developed long-term primary cultures of ependymal cells derived from ventricular walls of telencephalon and hypothalamus or choroidal cells (modified ependymal cells) derived from plexuses dissected out of fetal or newborn mouse or rat brains. Cultures were established in serum-supplemented or chemically defined media after seeding on serum-, fibronectin-, or collagen-laminin-coated plastic dishes or semipermeable inserts. To identify and characterize cell types growing in our cultures, we used morphological features provided by phase contrast, scanning, and transmission electron microscopy. We used antibodies against intermediate filament proteins (vimentin, glial fibrillary acidic protein, cytokeratin, desmin, neurofilament proteins), actin, myosin, ciliary rootlets, laminin, and fibronectin in single or double immunostaining, and monoclonal antibodies against epitopes of ependymal or endothelial cells, to recognize ventricular wall cell types with immunological criteria. Ciliated or nonciliated ependymal cells in telencephalic cultures, tanycytes and ciliated and nonciliated ependymal cells in hypothalamic cultures always exceeded 75% of the cultured cells under the conditions used. These cells were characterized by their cell shape and epithelial organization, by their apical differentiations observed by scanning and transmission electron microscopy, and by specific markers (e.g., glial fibrillary acidic protein, ciliary rootlet proteins, DARPP 32) detected by immunofluorescence. All these cultured ependymal cell types remarkably resembled in vivo ependymocytes in terms of molecular markers and ultrastructural features. Choroidal cells were also maintained for several weeks in culture, and abundantly expressed markers were detected in both choroidal tissue and culture (Na+-K+-dependent ATPase, DARPP 32, G proteins, ANP receptors). In this review, the culture models we developed (defined in terms of biological material, media, substrates, duration, and subculturing) are also compared with those developed by other investigators during the last 10 years. Focusing on morphological and functional approaches, we have shown that these culture models were suitable to investigate and provide new insights on (1) the gap junctional communication of ependymal, choroidal, and astroglial cells in long-term primary cultures by freeze-fracture or dye transfer of Lucifer Yellow CH after intracellular microinjection; (2) some ionic channels; (3) the hormone receptors to tri-iodothyronine or atrial natriuretic peptides; (4) the regulatory effect of tri-iodothyronine on glutamine synthetase expression; (5) the endocytosis and transcytosis of proteins; and (6) the morphogenetic effects of galactosyl-ceramide. We also discuss new insights provided by recent results reported on in vitro ependymal and choroidal expressions of neuropeptide-processing enzymes and neurosecretory proteins or choroidal expression of transferrin regulated through serotoninergic activation.

Choroidal readaptation to gravity in rats after spaceflight and head-down tilt.

To determine when choroidal structures were restored after readaptation to Earth gravity or orthostatic position, fine structure and protein distribution were studied in rat choroid plexus dissected either 6 h [Space Life Sciences-2 (SLS-2) experiments] or 2 days [National Institutes of Health-Rodent 1 (NIH-R1) experiments] after a spaceflight, or 6 h after head-down tilt (HDT) experiments. Apical alterations were noted in choroidal cells from SLS-2 and HDT animals, confirming that weightlessness impaired choroidal structures and functions. However, the presence of small apical microvilli and kinocilia and the absence of vesicle accumulations showed that the apical organization began to be restored rapidly after landing. Very enlarged apical microvilli appeared after 2 days on Earth, suggesting increased choroidal activity. However, as distributions of ezrin and carbonic anhydrase II remained altered in both flight and suspended animals after readaptation to Earth gravity, it was concluded that choroidal structures and functions were not completely restored, even after 2 days in Earth's gravity.

Superior cervical ganglionectomy suppresses circadian corticotropic rhythms in male rats in the short term (5 days) and long term (10 days).

Superior cervical ganglionectomy (SCGx) has drastic effects on numerous hormonal circadian rhythms and particularly on pineal melatonin secretion. We investigated the hormonal consequences of ablation of the superior cervical ganglion on the corticotropic circadian rhythms in the male rat. Plasma were obtained by sampling blood every 4 h, using a chronic carotid cannula. Adreno-corticotropin hormone (ACTH) was assayed by radioimmunoassay (RIA) and corticosterone (B) by radiocompetition. Urinary 6-sulphatoxymelatonin (aMT6s), considered as an index of the pineal gland activity, was assayed by specific RIA: a decrease in the aMT6s concentration after ganglionectomy was taken as proof of adequate surgical operation. Control animals showed classical circadian rhythms for ACTH and B with basal values during the light phase and circadian peaks around the light/dark interface. Five and ten days after ganglionectomy, the circadian rhythms of ACTH and B were suppressed. In addition, the mean ACTH concentrations increased significantly 10 days after ganglionectomy compared to those in sham-operated rats and 5 days post-operation group. The mean plasma corticosterone levels were similar in those three groups of animals. This is the first study demonstrating the suppressive effect of superior cervical ganglionectomy on the circadian corticotropic hormonal cycle.

Effects of acute tilt from orthostatic to head-down antiorthostatic restraint and of sustained restraint on the intra-cerebroventricular pressure in rats.

The tail-cast suspension rat model was developed to explore in ground laboratories the physiological effects of some of the stresses prevailing during space flight including and among them those of the headwards body fluid shifts. We recently showed in rats that an acute head-down tilt (45 degrees) from tail-cast orthostatic (OR) to antiorthostatic restraint (AOR) induced within 30 min and for 2 to 4 h an acute stress-like surge in plasma ACTH and corticosterone levels. Considering the proximity of the CRF producing neurons with the 3rd ventricle, we decided to explore the acute and longer-term effects of the OR/AOR tilt on the intra-cerebroventricular pressure (Picv) measured with an indwelling sensor-transmitter catheter stereotaxically implanted in the 3rd ventricle. At 1- or 10-min intervals the unit sent radiotelemetric signals for both Picv and motor activity (MA) to a receiver coupled with an automatic data analyser. The acute AOR-tilt induced within 10 min and for 60 min a 2.5-fold rise in Picv which receded to baseline between 60 and 90 min. During this time, the normally close correlation between Picv and MA was lost, as assessed by Spearman's rank coefficient. In a long-term experimental series we explored the evolution of both Picv and MA in individual rats subjected successively to a 7 day control phase (C). 7 days OR, and 3 days AOR. After the 1-h-long post-tilt rise of the Picv, the mean Picv levels measured for the next 3 days decreased significantly vs. both the preceding OR phase (-30%) and the initial C Phase (-40%). The circadian pattern of the diurnal Picv profile was impaired, as evidenced by a significant fall (i) in the night/day ratio (-25% vs. C). and (ii) even more in the spectral power of the circadian 1 c/24 h frequency (-85% vs. C). The simultaneously recorded MA fluctuations similarly displayed an altered diurnal pattern with a spectral power of the circadian frequency reduced to 7% of controls. However, contrary to the short-term experiment, in the long-term study the large alterations to both Picv and MA were strongly correlated, as during the control phase. The mechanisms involved in the swift post-tilt rise in the Picv together with an aroused corticotropic axis, and in the impact of sustained head-down restraint on CNS-controlled adaptive regulations including their circadian rhythms remain unknown.

Short-day stimulation of testicular activity and immunoreactivity of the hypothalamic GnRH system in mink following deafferentation of the pineal body by bilateral superior cervical ganglionectomy and melatonin replacement.

The effects of superior cervical ganglionectomy on testicular function (testis volume and plasma testosterone levels) and the immunocytochemical activity of the GnRH hypothalamic system were studied in the mink, a short-day breeder. Animals reared in a natural photoperiod were (i) ganglionectomized at four different times during the period extending from the end of summer to the end of autumn (September 15, October 20, October 28, and December 1), and (ii) reared for 50 days in a short gonadostimulatory photoperiod (4L:20D). Lastly, an attempt was made to overcome the effects of superior cervical ganglion removal by administering melatonin to mink reared in a natural photoperiod. In mink reared in a natural photoperiod, deafferentation of the pineal on September 15 (L = 12.5 h) or October 20 (L = 10.5 h) resulted in consistently low values of testicular volume and plasma testosterone until the end of the experiment (February). When the operation was performed on October 28 (L = 10 h) testicular activity was initiated but only lasted a short time and did not allow maximal gonadal development. When superior cervical ganglionectomy was carried out on December 1 (L = 8.5 h), during the phase of renewed testicular activity, the increases in testicular volume and testosterone levels were not affected by the operation and the subsequent variation of these parameters was identical to that observed in intact animals. Similarly, in mink reared for 50 days in a photoperiod of 4L:20D before superior cervical ganglionectomy, deafferentation of the pineal did not prevent gonadostimulation induced by short days.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in apical organization of choroidal cells in rats adapted to spaceflight or head-down tilt.

Structural changes observed in choroid plexuses from rats dissected aboard a space shuttle, on day 13 of an orbital flight (NASA STS-58 mission, SLS-2 Experiments) demonstrated that choroidal epithelial cells display a modified organization in a microgravitational environment. Results were compared with ultrastructural observations of choroid plexus from rats maintained under anti-orthostatic restraint (head-down tilt) for 14 days. In both experiment types, the main alterations observed by transmission electron microscopy, at the level of choroidal epithelial cells from the third and fourth ventricles, concerned the formation and the organization of apical microvilli, whereas pseudopod-like structures appeared. Immunocytochemical distribution of ezrin, a cytoskeletal protein involved in apical cell differentiation in choroid plexus, confirmed the structural alteration of microvilli in head-down tilted rats, Kinocilia tended to disappear from the apical surface, suggesting a partial loss of cell polarization. In addition, large amounts of clear vesicles were gathered in the apical cytoplasm of choroidal epithelial cells. Disorganization of apical microvilli accumulations of apical vesicles and partial loss of cell polarity showed that long-stays in weightlessness induced alterations in the fine structure of choroid plexus, consistent with a marked reduction of cerebrospinal fluid production.

Effects of an 11-day spaceflight on the choroid plexus of developing rats.

Cellular distributions of ezrin, a cytoskeletal protein involved in apical cell differentiation in choroid plexus, and carbonic anhydrase II, which is partly involved in the cerebrospinal fluid production, were studied by immunocytochemistry, at the level of choroidal epithelial cells from the lateral, third and fourth ventricles in normal or experimental fetuses, in parallel with the ultrastructure of apical microvilli, observed by transmission electron microscopy. We compared choroid plexuses from developing normal rats (gestational day 15 to birth) with choroid plexuses from 20-day-old rat fetuses, developed for 11 days in space, aboard a space shuttle (NASA STS-66 mission, NIH-R1 experiments), from gestational day 9 to day 20. The main changes observed in fetuses developed in space were demonstrated by immunocytochemistry and concerned the distribution of ezrin and carbonic anhydrase II. Thus, in fetuses developing in space, ezrin was strongly detected in the choroidal cytoplasm and weakly associated to the membrane in the apical domain of the choroid plexus from the fourth ventricle. Such alterations suggested that choroid plexus from rat fetal brain displays a delayed maturation under a micro-gravitational environment. In contrast, intense immunoreactions to anti-carbonic anhydrase II antibodies showed that this enzyme is very abundant in rats developed in space, compared to ground control fetuses.

Minor head injury at paediatric age in Argentina.

BACKGROUND: To evaluate epidemiologic factors in relation to paediatric head injury in the city of Mar del Plata, Argentina. METHODS: Retrospective (12-month) and prospective (6-month) evaluations were carried out on a total of 2,492 children with head injury (age range 0-15 years), taking into account sex, age, cause of injury, time of injury, incidence of hospitalisation, and the occurrence of fractures on conventional X-ray and computarized tomography (CT) scans of the head. Data were compiled and analyzed through a register called "Traumatic Brain Injury Register". RESULTS: From the total 2,492 children entered in the registry, 62% were males and 38% females. Age distribution was as follows: 33.5% from 0-2 years, 43.8% from 3-9 years, and 22.7% from 10-15 years. Falls were the main cause of head injury in children under 3 years of age, whereas car accidents prevailed among older groups (chi2 = 216.00, p<0.0001). Injuries occurred most frequently (84.7% of the cases) between 12:00 hrs and 24:00 hrs. From the total sample, 1,015 children (19.9%) required hospitalisation. Skull fractures were identified on conventional X-rays in 14.9% of the cases, and there was a direct relationship between skull X-rays and CT scans with respect to skull fractures (chi2 = 13.13, p<0.0001). The overall incidence of paediatric head injuries was 566 cases per 100,000 population and mortality rate was 4.65 per 1,000 head injuries. CONCLUSIONS: Our figures revealed an overall incidence of paediatric head injuries, which is 2.97 higher than the international average, suggesting that accident prevention measures should be further implemented.

Seasonal variations of pulsatile luteinizing hormone release in the mink (Mustela vison).

The pulsatile secretion of the hypophyseal luteinizing hormone (LH) is induced by the pulsatile secretion of the hypothalamic neurons secreting gonadotrophin-releasing hormone (GnRH). Seasonal variations in the pulsatility of LH were studied in the adult male mink (Mustela vison), reared under natural environmental conditions. Twenty-one animals were studied according to five critical phases in the breeding season: (1) the terminal phase of sexual quiescence, which precedes renewal of gonadal activity (October-November); (2) renewal of gonadal activity (December); (3) maximum gonadal activity at the height of the breeding season (February); (4) reduction of testicular activity (April); and (5) the initial phase of testicular quiescence (June). Levels of gonadal growth and activity were used to define each phase. A second animal group was studied after being reared for 2 months in an experimental gonado-inhibitory photoperiod, which, necessarily for the mink, was of the "long-day" type: 20L:4D regimen in the present study. Results, obtained with fully conscious animals, provide evidence for the pulsatile secretion of gonadotrophic hormone in this species. In spite of inter-individual differences in pulse patterns, particularly in phases 1 and 2, the pulsatile character of LH secretion is seen to vary markedly as a function of gonadal activity. The variations reflect an increase of hypophyseal activity as early as the preparative phase to the breeding season, and a decrease of activity during the testicular regression phase, which is followed by the onset of gonadotrophic quiescence in June. The main parameter affected statistically by these seasonal fluctuations is pulse frequency; variations in pulse frequency correlated with variations in mean plasma concentrations of LH. In the experimental gonado-inhibitory photoperiod, which led to a severe reduction in gonadal activity, all hormonal pulsatility parameters were statistically reduced; this confirms the importance of photoperiodic control of reproduction in Mustela vison. Several possible mechanisms are proposed for photoperiodic control.

Effects of bilateral olfactory bulbectomy on circadian rhythms of ACTH, corticosterone, motor activity and body temperature in male rats.

Bilateral olfactory bulbectomy (BOX) has major biochemical and behavioral effects, and is one of the most widely investigated of animal models of depression. We studied the consequences of BOX in male rats, on the organization of endogenous circadian rhythms for ACTH, corticosterone (Cort), motor activity (MA) and body temperature (BT). Mean levels were increased for Cort and MA, whereas no significant changes were observed for ACTH and BT. Significantly higher plasma Cort morning values were evidenced in BOX than sham-operated animals. In addition, compared with the single prominent power spectrum for the 24 hours period of control rats, the BOX animals displayed substantially lower 24 hours spectral power for the MA and BT circadian rhythms. These alterations suggest that olfactory bulbectomy, by disruption of the afferences and efferences, induced drastic changes in the function of the endogenous clock or of its regulating systems. From this point of view, bulbectomized rats may therefore be a valuable model to studying the etiology of psychiatric disorders with rhythm disturbance.