Active zone proteins are dynamically associated with synaptic ribbons in rat pinealocytes.

Synaptic ribbons (SRs) are prominent organelles that are abundant in the ribbon synapses of sensory neurons where they represent a specialization of the cytomatrix at the active zone (CAZ). SRs occur not only in neurons, but also in neuroendocrine pinealocytes where their function is still obscure. In this study, we report that pinealocyte SRs are associated with CAZ proteins such as Bassoon, Piccolo, CtBP1, Munc13-1, and the motorprotein KIF3A and, therefore, consist of a protein complex that resembles the ribbon complex of retinal and other sensory ribbon synapses. The pinealocyte ribbon complex is biochemically dynamic. Its protein composition changes in favor of Bassoon, Piccolo, and Munc13-1 at night and in favor of KIF3A during the day, whereas CtBP1 is equally present during the night and day. The diurnal dynamics of the ribbon complex persist under constant darkness and decrease after stimulus deprivation of the pineal gland by constant light. Our findings indicate that neuroendocrine pinealocytes possess a protein complex that resembles the CAZ of ribbon synapses in sensory organs and whose dynamics are under circadian regulation.

The "Mainzer EMF-Wachhund": results from a watchdog project on self-reported health complaints attributed to exposure to electromagnetic fields.

The "Mainzer EMF-Wachhund," a watchdog project, offered a system of self-notification of health complaints attributed to exposures to electromagnetic fields (EMFs) to a population of a part of Germany with about 4 million inhabitants. By using a self-administered questionnaire, which was provided online and for download from the Internet, 192 persons reported such health complaints in the period from October 2003 to March 2005. Of these, 56% classified themselves as electromagnetic hypersensitive (EH). Predictors of this self classification were being affected by all kinds of EMF rather than single EMF sources and being female. On average, EH subjects reported a high degree of suffering, 77% of whom had already sought advice from physicians. An Internet-based standardized questionnaire is an economic way of offering affected persons a direct link to scientific institutions to establish contact. However, the study base obtained by such an approach is not representative to estimate a population-based prevalence. As a large number of subjects did not classify themselves as EH and reported very specific links between exposure and symptoms, they may provide a very distinct and interesting group for future research.

Circadian gene expression patterns of melanopsin and pinopsin in the chick pineal gland.

The directly light-sensitive chick pineal gland contains at least two photopigments. Pinopsin seems to mediate the acute inhibitory effect of light on melatonin synthesis, whereas melanopsin may act by phase-shifting the intrapineal circadian clock. In the present study we have investigated, by means of quantitative RT-PCR, the daily rhythm of photopigment gene expression as monitored by mRNA levels. Under a 12-h light/12-h dark cycle, the mRNA levels of both pigments were 5-fold higher in the transitional phase from light to dark than at night, both in vivo and in vitro. Under constant darkness in vivo and in vitro, the peak of pinopsin mRNA levels was attenuated, whereas that of melanopsin was not. Thus, whereas the daily rhythm of pinopsin gene expression is dually regulated by light plus the intrapineal circadian oscillator, that of melanopsin appears to depend solely on the oscillator.

Arylalkylamine N-acetyltransferase gene expression in retina and pineal gland of rats under various photoperiods.

The present study examines how the circadian oscillators in the retina and the suprachiasmatic nucleus (SCN) respond to changes in photoperiod. Arylalkylamine N-acetyltransferase (aa-nat) gene expression studied by quantitative RT-PCR revealed that in adult Sprague-Dawley rats kept under different light-dark (LD) cycles for two weeks the temporal pattern of AA-NAT mRNA expression was identical in retina and pineal gland. In both tissues, the time span between the onset of darkness and the nocturnal rise in AA-NAT mRNA expression was 3 h under LD 20:4, 6 h under LD 12:12, and 15 h under LD 4:20. As aa-nat expression in the pineal gland is regulated by the circadian oscillator in SCN, the results suggest that the photoperiodic differences accompanying the seasons of the year are imprinted in more than one oscillator and that this may accentuate the important message regarding 'time of year.'

Mouse photoreceptor synaptic ribbons lose and regain material in response to illumination changes.

Abstract Chemical synapses equipped with ribbons are tonically active, high-output synapses. The ribbons may play a role in the trafficking of synaptic vesicles. Recent findings in retinal rod cells of BALB/c mice indicate that ribbons are large and smooth in the dark phase, and, due to the formation and release of protrusions, small during the light phase. As a consequence of these changes, ribbons may traffick fewer vesicles in the light than in the dark phases. The aim of the present study was to find out whether the above ribbon changes in this mouse strain are strictly illumination-dependent and which signalling processes may be involved. Here, we show that ribbons form protrusions and release them into the cytoplasm within 30-60 min after lights on, the reverse occurring within 30 min after lights off. Under constant light or constant dark, no circadian rhythm of synaptic ribbon changes is observed. The illumination-dependence of ribbon structure is supported by in vitro experiments showing that in dark-adapted retinas, light induces the same morphological changes as in vivo. In vitro, the effect of light on the ribbons can be counteracted by cyclic guanosine monophosphate and melatonin. In dark-adapted retinas, light effects can be produced by decreasing the calcium ion concentrations in the incubation media. These results suggest that in retinal rod cells, the well known phototransduction signalling mechanisms may be responsible for the ribbon changes presently and previously reported.

Rat pineal arylalkylamine-N-acetyltransferase: cyclic AMP inducibility of its gene depends on prior entrained photoperiod.

The nocturnal biosynthesis of melatonin in the rat pineal depends on strongly enhanced expression of the enzyme N-acetyltransferase [arylalkylamine N-acetyltransferase (AA-NAT); EC 2.3.1.87]. AA-NAT transcription is stimulated during darkness by adrenergic inputs to the pineal from the suprachiasmatic nucleus (SCN). Nocturnal activation of the AA-NAT promotor following stimulation of pinealocyte adrenoceptors involves cAMP-dependent stimulation of protein kinase A (PKA). The nocturnal rise in AA-NAT depends on the lighting conditions. As compared with light/dark (LD) 12:12, the delay between dark onset and the nocturnal rise in AA-NAT is shortened under long photoperiods and prolonged under short photoperiods. Here, we report that the rapidity of nocturnal AA-NAT induction depends on cAMP inducibility of the gene. Accordingly, cAMP produces a strong AA-NAT response in pineals obtained from rats housed under long photoperiods and a weak AA-NAT response under short photoperiods. Changes in AA-NAT inducibility are fully developed not earlier than after seven cycles. This observation suggests that long-term changes in the photoperiod are necessary to achieve full adjustment of cAMP inducibility of the gene. A direct relationship was found between cAMP-dependent AA-NAT inducibility and the pineal protein kinase A (PKA) activity. As compared to LD 12:12, PKA activity was increased under LD 20:4 and attenuated under LD 4:20. On the basis of the present findings, we suggest that the photoperiod determines the effectiveness of nocturnal AA-NAT induction by long-term modulation of the intrapineal pathway that transmits the cAMP signal to the AA-NAT gene.

The phototransduction cascade in the isolated chick pineal gland revisited.

It is well established that the isolated chick pineal gland is directly light sensitive and that melatonin synthesis of the gland can be inhibited by exposing the gland to light during scotophase. Since not all the steps of the phototransduction cascade have been clarified to the same extent as in the retina, we have treated isolated chick pineal glands with 90 min of light during scotophase and with drugs that affect key-components of vertebrate phototransduction, i.e., cyclic guanosine monophosphate (cGMP) phosphodiesterase 6 (PDE6), cGMP levels and cGMP-gated calcium channels. The endpoint measured was the activity of the rate-limiting enzyme of melatonin synthesis, arylalkylamine N-acetyltransferase (AA-NAT), which is inhibited by light. The effects on AA-NAT activity of light were negated by addition of dipyridamol and zaprinast, either of which inhibits the light-induced activation of PDE6. The effect of light was also counteracted by the nitric oxide donor sodium nitroprusside and C-type natriuretic peptide, both of which increase cGMP levels, and by the calcium channel agonist Bay K 8644, which prevents the cGMP-decrease-induced closure of cGMP-gated calcium channels. Inhibition of nitric oxide synthase (NOS) by N(G)-nitro-l-arginine did not influence the inhibitory effect of light, suggesting that the NOS pathway does not play a role. Since the light effect on AA-NAT activity involves both cGMP and cyclic adenosine monophosphate (cAMP) hydrolysis, we have also studied whether the cGMP-inhibited cAMP phosphodiesterase 3 (PDE3) is involved. As the specific PDE3 inhibitor cilostamide is without effect, we assume that the light-induced decrease of cAMP levels does not involve PDE3. These results taken together strongly suggest that the investigated steps of the phototransduction cascade in the isolated chick pineal gland are basically similar to those in the retina.

Plasticity of synaptic ribbons of the rat pineal gland in vitro--minor effects of electrical stimulation.

Synaptic ribbons (SRs) of mammalian pinealocytes exhibit day/night changes in number and size, changes that are apparently regulated by the suprachiasmatic nucleus via postganglionic sympathetic nerve fibres. Since the neural control of SR changes is far from clear and as pinealocytes produce action potentials, we undertook to investigate whether electrical stimulation affects SR changes. Isolated rat pineal glands removed during the daytime were kept in vitro for 0, 30, 60, 90 or 120 min, with or without continuous electrical stimulation (1 mA, 1 Hz), followed by the quantification of SR profiles (SRPs) by transmission electron microscopy. SRs were categorised as to whether they lay less than 100 nm away from the pinealocyte plasmalemma (SRPs(near)) or more distant from it (SRPs(dist)) and the lengths of the profiles were measured. Cultured pineal organs showed a significant numerical depression of SRPs(near), irrespective of whether the organs had been electrically stimulated or not. SRPs(near) length revealed a significant increase at 60 min in unstimulated control tissue and at 30 min in electrically stimulated glands. SRPs(dist) length decreased significantly at 30 min in control glands and after 60 min in electrically stimulated glands. Thus, action potentials inside the pineal gland appear to be minor factors regulating SR numbers. In future pineal studies, SRPs(near) and SRPs(dist) should be considered separately as they differ in plasticity.

Teaching and learning gross anatomy using modern electronic media based on the visible human project.

This article reviews online (Internet) anatomy projects and multimedia productions (books and CD-ROMs) based on the Visible Human Project (VHP) of the United States National Library of Medicine. The focus of these projects and productions is on the teaching of 3D anatomy using the digitized sections of the visible human male and female. The article also provides information about the VHP, its goals and what it has achieved thus far.

[Chrono-endocrinology--quo vadis?]. / Chronoendokrinologia--quo vadis?

The present review deals with important new chronobiological results especially in the field of chronoendocrinology, shedding new light on the circadian organisation of mammals including man. In vitro studies have shown that the concept of the existence of a single circadian oscillator located in the suprachiasmatic nucleus has to be extended. Circadian oscillators have also been found to exist in the retina, islets of Langerhans, liver, lung, and fibroblasts. Another major result is the detection of a new photopigment, melanopsin, present in a subpopulation of retinal ganglion cells which are lightsensitive and project to the suprachiasmatic nucleus, acting as zeitgeber for the photic entrainment of the circadian rhythm. We are only beginning to understand how the circadian oscillator transmits the circadian message to the endocrine system. The generation of circadian and seasonal rhythms of hormone synthesis is best understood in the pineal gland and its hormone melatonin. Seasonal changes of melatonin synthesis are transduced in the pars tuberalis of the adenohypophysis which is now entering the limelight of chronoendocrinological research. Currently, the elucidation of the genetic basis and the molecular organisation of the circadian oscillator within individual cells is a major thrust in chronobiological research.

Anatomy online: presentation of a detailed WWW atlas of human gross anatomy--reference for medical education.

We present an online anatomy atlas based on the Visible Human Project (VHP) of the US National Library of Medicine. The objective is to provide original unlabeled as well as labeled sections of the human body of high quality and resolution on the Internet, for use in basic and continuing medical education. For a representative overview of the body, 370 axial sections were selected from the male and female data base of the VHP with special regard to regions of clinical interest. Each section is accompanied by its corresponding computer tomography (CT) image and, if available, magnetic resonance images (MRI) for quick and easy comparison of morphologic and radiologic structures. The sections can be studied unlabeled or labeled according to the current Terminologia Anatomica. A linked vocabulary with more than 850 terms explains the labeling. Animations of the sections as well as of CT and MR images allow for further visualization of the topographic relationships of anatomical structures. The responses to the project indicate that students and physicians regard the Internet Atlas of Human Gross Anatomy as a most useful aid for learning and reviewing anatomical details. The atlas is accessible on: http://www.uni-mainz.de/FB/Medizin/Anatomie/workshop/vishuman/Eready.html.