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Whereas short-term plasticity is often initiated on one side of the synapse, long-term plasticity involves coordinated changes on both sides, implying extracellular signaling. We have investigated the possible signaling role of an Aplysia neurotrophin (ApNT) in facilitation induced by serotonin (5HT) at sensory-to-motor neuron synapses in culture. ApNT is an ortholog of mammalian BDNF, which has been reported to act as either an anterograde, retrograde, or autocrine signal, so that its pre- and postsynaptic sources and targets remain unclear. We now report that ApNT acts as a presynaptic autocrine signal that forms part of a positive feedback loop with ApTrk and PKA. That loop stimulates spontaneous transmitter release, which recruits postsynaptic mechanisms, and presynaptic protein synthesis during the transition from short- to intermediate-term facilitation and may also initiate gene regulation to trigger the transition to long-term facilitation. These results suggest that a presynaptic ApNT feedback loop plays several key roles during consolidation of learning-related synaptic plasticity.

Whereas short-term synaptic plasticity is often either pre- or postsynaptic, intermediate- and long-term plasticity generally require coordinated pre- and postsynaptic mechanisms. Thus, the transition from presynaptic short-term facilitation (STF) to intermediate-term facilitation (ITF) induced by 5HT at Aplysia sensory-to-motor neuron synapses requires the recruitment of postsynaptic mechanisms and activation of protein synthesis in both neurons. In the companion paper to this report, we found that presynaptic autocrine signaling by an Aplysia neurotrophin (ApNT) forms a positive feedback loop that drives the synapses from STF to ITF. Here we report that ApNT also acts through both anterograde and retrograde signaling to form a transsynaptic positive feedback loop that orchestrates cellular functions in both the presynaptic and postsynaptic neurons during the induction of ITF. These two feedback loops activate protein synthesis in each synaptic compartment, which in both cases depends on signaling from the other synaptic compartment. These results suggest that the pre- and postsynaptic compartments act as one functional unit during the consolidation of learning-related facilitation induced by 5HT.

Whereas short-term synaptic plasticity is often either pre- or postsynaptic, intermediate- and long-term plasticity generally require coordinated pre- and postsynaptic mechanisms. Thus, the transition from presynaptic short-term facilitation (STF) to intermediate-term facilitation (ITF) induced by 5HT at Aplysia sensory-to-motor neuron synapses requires the recruitment of postsynaptic mechanisms and activation of protein synthesis in both neurons. In the companion paper to this report, we found that presynaptic autocrine signaling by an Aplysia neurotrophin (ApNT) forms a positive feedback loop that drives the synapses from STF to ITF. Here we report that ApNT also acts through both anterograde and retrograde signaling to form a transsynaptic positive feedback loop that orchestrates cellular functions in both the presynaptic and postsynaptic neurons during the induction of ITF. These two feedback loops activate protein synthesis in each synaptic compartment, which in both cases depends on signaling from the other synaptic compartment. These results suggest that the pre- and postsynaptic compartments act as one functional unit during the consolidation of learning-related facilitation induced by 5HT.

Environmental factors during early life stages affect behavioral and physiological phenotypes in adulthood. We examined the effect of photoperiods during development on neurogenesis and affective behaviors during adolescence/adulthood using C57BL/6J mice. Mice were born and raised until weaning under long-day conditions (LDs) or short-day conditions (SDs), followed by a 12L12D cycle until adulthood. Adult mice born under SD showed a shorter latency to first immobility in the forced swim test when compared with the mice born under LD. The mice born under SD also exhibited significantly lower prepulse inhibition, which is a characteristic of schizophrenia. However, the mice exposed to SD and LD during the prenatal period only did not show differences in prepulse inhibition. At 4 weeks of age, there were less 5-bromo-2′-deoxyuridine (BrdU)-positive cells in the dentate gyrus (DG) of the hippocampus of mice born under SD when compared with mice born under LD. Double immunostaining showed that the mice born under SD showed less BrdU/glial fibrillary acidic protein (GFAP, an astrocyte marker) cells when compared with mice born under LD. Furthermore, expression of the glucocorticoid receptor in the DG was higher in mice born under SD, and the photoperiod-dependent changes in the number of BrdU-positive cells in the DG were abolished by administration of RU486, a glucocorticoid receptor antagonist. These results suggest that the photoperiod in early life alters astrogenesis in the hippocampus via the hypothalamic–pituitary-adrenal axis and may relate to affective behaviors in adulthood.

Throughout life, we are exposed to a variety of stresses, which may be inevitable and noxious sometimes. During evolution, animals must have acquired some physiological means to counteract stress. Vascular endothelial growth factor (VEGF) is an angiogenic and neurogenic factor, which has been shown to elicit antidepressant-like effects in response to different external stimuli, potentially functioning as an anti-stress molecule. However, it remains largely unknown how VEGF modulates mood-related behaviors. To investigate molecular correlates, we analyzed monoaminergic systems of VEGF transgenic mice that display antidepressant-like behavior. Immunostaining showed that overall morphologies of monoaminergic nuclei and their processes were normal. However, we found imbalances in brain monoamine contents, in which the levels of norepinephrine and serotonin, but not dopamine, were decreased exclusively in the regions where VEGF was expressed. The turnover of norepinephrine showed a marked increase and serotonin turnover showed a modest reduction, whereas dopamine turnover was not affected. The protein levels of tyrosine hydroxylase and tryptophan hydroxylase, the rate-limiting enzymes of catecholamine and serotonin synthesis, remained constant. The mRNA levels of monoamine receptors were generally similar but adrenergic receptors of ADRα1A and ADRβ1 were down-regulated. Behavioral tests showed that serotonin- or norepinephrine-selective antidepressant drugs failed to additively enhance antidepressant-like behaviors, whereas monoamine depleting drugs attenuated VEGF-mediated antidepressant-like effect. These data suggest that VEGF-induced antidepressant-like effects involve modulation of norepinephrine and serotonin systems.

Visual perception is important for acquiring spatial information in many animals, and loss of vision often causes devastating effects on their survival. However, it may be compensated to some extent by utilizing other intact sensory modalities. The cone-rod homeobox (Crx) gene plays a key role in development of photoreceptor cells, but behavioral consequences of the gene deletion have not been well characterized. In this study, we analyzed homozygous knockout (Crx(-/-)) mice by comparing with heterozygous knockout (Crx(+/-)) mice as controls. We first checked their vision with three different behavioral paradigms of the glass table visual recognition test, the light-dark transition test, and the Barnes maze test with a visual cue, all of which indicated that Crx(-/-) mice were blind while Crx(+/-) mice were sighted. In the fear conditioning test, Crx(-/-) mice were able to acquire both contextual and cued memory using non-visual information. Crx(-/-) mice showed normal thigmotaxis, but the exploratory activities were significantly increased. In the elevated plus maze test, it was unexpected that Crx(-/-) mice rarely fell down from the narrow platform. There was no reduction in their moving speeds and the moving distance was rather increased in Crx(-/-) mice. Such behaviors were not affected by trimming their whiskers. However, attachment of earplugs significantly reduced their exploratory activities. In summary, these data suggest that Crx(-/-) mice were behaviorally blind but were able to learn and recognize external environment utilizing non-visual information, as exemplified by sensory compensation in exploratory activities on elevated platforms.

The present study examined the effects of enzymatically hydrolyzed collagen peptides on the level of hippocampal neurogenesis and emotional behavior in adult mice. For this purpose, two kinds of enzymatically hydrolyzed collagen peptides, the lower or higher molecular weight peptides (LP: below 2,000, HP: about 30,000) were administered orally to C57BL/6 mice for 4 weeks. As a result, the density of proliferating cells in subgranular zone of hippocampus showed a 1.2-fold increase in LP mice as compared with HP mice. Additionally, LP mice spent less time in closed arms than HP mice in elevated plus maze test to examine anxiety-related behavior. These results suggest that oral administration of the lower molecular weight peptides derived from collagen enhanced the hippocampal neurogenesis and exerted emotional behavior in adult mice.

Imaging studies have shown that even the earliest phases of long-term plasticity are accompanied by the rapid recruitment of synaptic components, which generally requires actin polymerization and may be one of the first steps in a program that can lead to the formation of new stable synapses during late-phase plasticity. However, most of those results come from studies of long-term potentiation in rodent hippocampus and might not generalize to other forms of synaptic plasticity or plasticity in other brain areas and species. For example, recruitment of presynaptic proteins during long-term facilitation by 5HT in Aplysia is delayed for several hours, suggesting that whereas activity-dependent forms of plasticity, such as long-term potentiation, involve rapid recruitment of presynaptic proteins, neuromodulatory forms of plasticity, such as facilitation by 5HT, involve more delayed recruitment. To begin to explore this hypothesis, we examined an activity-dependent form of plasticity, homosynaptic potentiation produced by tetanic stimulation of the presynaptic neuron in Aplysia . We found that homosynaptic potentiation involves presynaptic but not postsynaptic actin and a rapid (under 10 min) increase in the number of clusters of the presynaptic vesicle-associated protein synaptophysin. These results indicate that rapid recruitment of synaptic components is not limited to hippocampal potentiation and support the hypothesis that activity-dependent types of plasticity involve rapid recruitment of presynaptic proteins, whereas neuromodulatory types of plasticity involve more delayed recruitment.

Mandarins are picked by hand with the use of shears. An average mandarin picker makes at least 20,000 cuts daily. This daily workload on the upper limbs is the cause of occupational cervicobrachial disorders (OCD). In order to improve their working conditions, we devised two types of shears, one with a soft grip wrapped around the handle and another with shock reducing cushions in between the handles. The following two experiments were conducted. First we tested and compared the Borg scale and pinch meter scores of each participant by having them cut sticks as the same radius as those of mandarin stems using both the improved and conventional shears. Next, we placed two pressure sensors on each shear, where the middle finger and thumb contact the handles, to measure and compare the impact pressure within the hand. The results of the first experiment suggested that the improved shears reduces workload to the hand and fingers when comparing results to those of the conventional shears. The second experiment indicates that the shock reducing cushions reduces impact pressure to the hand from the handles when cutting.

By the two hybrid screening of mouse brain cDNA library, we identified Hppi, a cell death-promoting protein, as a binding partner of postsynaptic scaffold protein Homer1c. Hippi interacted specifically with Homer1c but not with its homologue Homer2. It was reported that Hippi, when complexed with Hip1, induces the apoptosis in striatal neurons and may cause Huntington’s disease. We found that this apoptotic effect of Hippi was specific to the striatum and was not observed in hippocampal neurons. Furthermore, the apoptotic effect of Hippi was prevented when Homer1c was co-expressed in cultured striatal neurons. The protective effect of Homer1c was diminished when Hippi binding domain was deleted. These results suggest that Homer1c may play an important role in the mechanisms of neuronal death in the striatum.

The number of nursing home has increased largely in Japan since 1990. The Long-term Care Insurance in 2000 has accelerated the increase of nursing homes. The care giving and cooking in nursing homes have high risk factors of muscle-skeletal diseases (MSDs). However, the working conditions have not yet been improved. Thus, the incidence of low back pain and cervico-brachial disorder is very high among the care workers and cooks. Therefore, it is important to prevent the MSDs among the care workers and cooks. This study has been conducted to make a model of the participatory improvement focusing on low back pain in a nursing home for three years. As a result of the study, many improvements have been implemented and the incidence of low back pain has been reduced.

Rho GTPases are key transducers of integrin/extracellular matrix and growth factor signaling. Although integrin-mediated adhesion and trophic support suppress neuronal apoptosis, the role of Rho GTPases in neuronal survival is unclear. Here, we have identified Rac as a critical pro-survival GTPase in cerebellar granule neurons (CGNs) and elucidated a death pathway triggered by its inactivation. GTP-loading of Rac1 was maintained in CGNs by integrin-mediated (RGD-dependent) cell attachment and trophic support. Clostridium difficile toxin B (ToxB), a specific Rho family inhibitor, induced a selective caspase-mediated degradation of Rac1 without affecting RhoA or Cdc42 protein levels. Both ToxB and dominant-negative N17Rac1 elicited CGN apoptosis, characterized by cytochrome c release and activation of caspase-9 and -3, whereas dominant-negative N19RhoA or N17Cdc42 did not cause significant cell death. ToxB stimulated mitochondrial translocation and conformational activation of Bax, c-Jun activation, and induction of the BH3-only protein Bim. Similarly, c-Jun activation and Bim induction were observed with N17Rac1. A c-jun N-terminal protein kinase (JNK)/p38 inhibitor, SB203580, and a JNK-specific inhibitor, SP600125, significantly decreased ToxB-induced Bim expression and blunted each subsequent step of the apoptotic cascade. These results indicate that Rac acts downstream of integrins and growth factors to promote neuronal survival by repressing c-Jun/Bim-mediated mitochondrial apoptosis.

Homer family proteins are encoded by three genes, homer1, 2 and 3. Most of these proteins are expressed constitutively in nervous systems and accumulated in postsynaptic regions. However, the functional significance of these proteins, especially the significance of the distinction among the proteins encoded by homer1, 2 and 3, is still obscure. In the present study, we isolated a cDNA clone encoding a novel protein by two-hybrid system screening using the C-terminal half of Homer2b as the bait. This protein, termed 2B28, has 297 amino acid residues and contains three major domains: a UBA domain, a coiled-coil region, and a UBX domain. When expressed in HEK293T cells, 2B28 showed colocalization with uniquitin and enhanced the expression levels of IkappaB or Homer1a proteins, which are known to be degraded by proteasomes, indicating that 2B28 is involved in ubiquitin-proteasome functions. 2B28 specifically interacted and colocalized with Homer2 proteins, but not with Homer1 proteins. So far, we have identified no counterpart of 2B28 for Homer1 experimentally or in the protein databases. These results suggest that the specific interaction of 2B28 with Homer2 may play a role in regulation of protein degradation by ubiquitin-proteasome systems and that this function may be specific to Homer2 proteins among Homer family proteins.

Summary Application of Clostridium difficile toxin B, an inhibitor of the Rho family of GTPases, at the Aplysia sensory to motor neuron synapse blocks long-term facilitation and the associated growth of new sensory neuron varicosities induced by repeated pulses of serotonin (5-HT). We have isolated cDNAs encoding Aplysia Rho , Rac , and Cdc42 and found that Rho and Rac had no effect but that overexpression in sensory neurons of a dominant-negative mutant of ApCdc42 or the CRIB domains of its downstream effectors PAK and N-WASP selectively reduces the long-term changes in synaptic strength and structure. FRET analysis indicates that 5-HT activates ApCdc42 in a subset of varicosities contacting the postsynaptic motor neuron and that this activation is dependent on the PI3K and PLC signaling pathways. The 5-HT-induced activation of ApCdc42 initiates reorganization of the presynaptic actin network leading to the outgrowth of filopodia, some of which are morphological precursors for the learning-related formation of new sensory neuron varicosities.

SummaryRecent results suggest that long-lasting potentiation at hippocampal synapses involves the rapid formation of clusters or puncta of presynaptic as well as postsynaptic proteins, both of which are blocked by antagonists of NMDA receptors and an inhibitor of actin polymerization. We have investigated whether the increase in puncta involves retrograde signaling through the NO-cGMP-cGK pathway and also examined the possible roles of two classes of molecules that regulate the actin cytoskeleton: Ena/VASP proteins and Rho GTPases. Our results suggest that NO, cGMP, cGK, actin, and Rho GTPases including RhoA play important roles in the potentiation and act directly in both the presynaptic and postsynaptic neurons, where they contribute to the increase in puncta of synaptic proteins. cGK phosphorylates synaptic VASP during the potentiation, whereas Rho GTPases act both in parallel and upstream of cGMP, in part by maintaining the synaptic localization of soluble guanylyl cyclase.

In cultured rat hippocampal neurons, overexpression of Homer1a/Vesl-1S, an inducible protein upregulated by seizure or long-term potentiation, caused a reduction of punctate distribution of a postsynaptic protein Homer1c/Vesl-1L, without significant decrease in its total amount. Clusters of F-actin were also decreased. Treatments of cells with BDNF or a proteasome inhibitor, which cause increase in the expression level of endogenous Homer1a, also resulted in the reduction of Homer1c puncta. These results indicate that the accumulation of Homer1a, either exogenously expressed or endogenously induced, caused redistribution and dispersion of postsynaptic clusters of Homer1c and F-actin, suggesting an important role of Homer1a in synaptic remodeling.

The time course and functional significance of the structural changes associated with long-term facilitation of Aplysia sensory to motor neuron synaptic connections in culture were examined by time-lapse confocal imaging of individual sensory neuron varicosities labeled with three different fluorescent markers: the whole-cell marker Alexa-594 and two presynaptic marker proteins—synaptophysin-eGFP to monitor changes in synaptic vesicle distribution and synapto-PHluorin to monitor active transmitter release sites. Repeated pulses of serotonin induce two temporally, morphologically, and molecularly distinct presynaptic changes: (1) a rapid activation of silent presynaptic terminals by filling of preexisting empty varicosities with synaptic vesicles, which parallels intermediate-term facilitation, is completed within 3–6 hr and requires translation but not transcription and (2) a slower generation of new functional varicosities which occurs between 12–18 hr and requires transcription and translation. Enrichment of empty varicosities with synaptophysin accounts for 32% of the newly activated synapses at 24 hr, whereas newly formed varicosities account for 68%.

We have developed a new armrest (NA) with adequate width supporting forearms, inclining upper surface, an area for mouse, and fixers at the edge of a shallow desk. We compared the cervicobrachial load among NA, NA with a mouse-pad (NAM) and a conventional palmrest with a mouse-pad (PRM) during word processing work. Seventeen healthy subjects operated a word processor for 1-h, alternately using 3 devices. EMG and subjective pain scores (PS) were recorded mainly from the right-side upper limb. EMG of the trapezius in PRM was significantly lower than in NAM; EMG of the anterior deltoideus (DELTa) in NA lower than in NAM; EMG of the medial deltoideus (DELTm) in NA lower than in PRM. The shoulder PS in NA/NAM was significantly lower than in PRM; the upper arm PS in NAM lower than in PRM; the forearm PS in NA/NAM lower than in PRM. These results suggest that NA reduces the cervicobrachial load than PRM.

An ergonomic measure can be effective in the treatment of musculo-skeletal disorders such as low back pain and cervicobrachial disorders. However, they have not yet been developed or implemented to their full potential at workplaces. One reason for this is that workers are rarely given the opportunity to be trained for ergonomic methods of improvement or to put these methods into practice in the work environment. An industrial medical doctor (industrial doctor), whose duties include planning and implementing ergonomic measures, as is the case in Japan, can play an important role in encouraging such improvements in the workplace. The purpose of this paper is to outline the role of the industrial doctor in planning and implementing ergonomic measures for reducing low back pain, and in fostering worker participation in the improvement process. A case study involving the maintenance division of a steel mill is used to illustrate the role of the industrial doctor. The role is summarized as follows: (1) Educate workers on the main causes of musculo-skeletal disorders in the workplace. (2) Propose to workers practical ergonomic principles to be implemented. (3) Propose participatory improvement methods to workers. (4) Propose to workers practical, simple and low-cost measures including supports. (5) Continuously restate and reinforce the goal of the ergonomic measures to workers. (6) Advise managers to cooperate in the process and budgeting. Relevance to industry This study is related to developing practical methods for ergonomic improvements in the workplace.

Recently there has been an increasing incidence of occupational cervicobrachial disorders (OCD) and writer's cramp in office workers using ballpoint pens in writing operations. For the sake of workers who use ballpoint pens, it is essential to prevent such health hazards. It has been observed that a strong gripping pressure on the ballpoint pen significantly contributes to the development of these conditions. The present authors have been developing a new ballpoint pen by altering the grip area in such a way as to reduce the gripping pressure, and thus prevent OCD. The purpose of this study is to compare our ballpoint pen (new pen) with a conventional ballpoint pen (conventional pen) for the load that they exert on the upper limb during one hour of continuous writing. Electromyograms (EMG) and upper limb pain scores are used as indicators. The conventional pen used was selected from commercially available ballpoint pens widely used in offices. The grip area is cylindrical with an 8.3 mm diameter. It is manufactured of hard plastic, which can make it feel rigid and slippery to the user. The new pen has a cylindrical grip area that flares out at the bottom, near the pen-tip, and has a diameter ranging from 11.9-13.6 mm. In addition, the grip is constructed of a 2 to 3 mm-thick silicon rubber sleeve that is softer and less slippery in comparison with the conventional pen. Twelve students (5 males and 7 females) without any preexisting cervicobrachial disorders were asked to transcribe an English text for one hour, alternately using the two kinds of pens. The EMG of the flexor pollicis brevis was measured and recorded every second, while subjective pain scores were recorded every five minutes for the thumb, forefinger, middle finger, thenar, forearm extensor (forearm) and shoulder. The EMG of the flexor pollicis brevis and the pain scores for the thumb, forefinger, middle finger, forearm and shoulder were significantly lower for the new pen than for the conventional pen. These results suggest that after an hour of continuous writing, the new pen reduces the muscle load on the upper limb, and therefore mitigates fatigue in this area.

Low back pain is a significant health problem for professional drivers. Therefore it is important to design a car driver seat with less load to reduce low back pain. The purpose of this study is to compare the low back load in a long-term driving experiment between two different types of driver seats (the new seat and the standard seat). The new seat has the following characteristics vs. the standard seat: (1) a lumbar support, (2) side supports, (3) seat is firmer at the ischial tuberosity region, but it is softer at the femoral region, (4) a head rest which is adjustable sagittally, and (5) a rubber mat under the seat. Sixty male subjects, who commuted 5 times/week for about 50 min/trip with the standard seat and complained of low back pain in driving, were selected from a group of 1600 subjects. The existing seat in their personal vehicles was replaced with the new seat for 30 subjects. For 6 months, 30 used the new seat, and the other 30 used the standard seat. Before, and 3 and 6 months after the start of the study, the low back load was estimated by lumbar examination and subjective evaluation of low back pain. At the second examination, the subjective evaluation significantly improved for the new seat (vs. the standard seat). At the third examination, the kinetic pain score in bending backward and the Lasegue test significantly improved for the new seat (vs. the standard seat). It is suggested that the low back load of the new seat was less than the standard seat. The factors of the lumbar support, the side supports, the firmer seat at the ischial tuberosity region, the adjustable head rest and the rubber mat all contributed to a reduction in low back load.

Pkn2, a protein Ser/Thr kinase, from the developmental bacterium Myxococcus xanthus was expressed under a T7 promoter in Escherichia coli and purified. Purified Pkn2 retained the autophosphorylation activity with the Km value of 177 microM for ATP and 73 nmol/min/mg for Vmax. The optimum pH and temperature were determined to be 7.5 and 35 degrees C, respectively. The autophosphorylation activity was inhibited by staurosporine with the IC50 value of 400 nM while H-7 and genistein had little effect on this kinase. Pkn2 appears to be unique for its higher manganese dependence. This is the first biochemical characterization of the prokaryotic protein Ser/Thr kinase.

Back support belts are one of the important measures for preventing low back pain. One theory attributes the effectiveness of such a belt to the intraabdominal pressure it applies. The purpose of this study was to investigate the effect of a pelvic belt on abdominal pressure when the subject held four different weights at four different bending angles. The subjects of the study were ten male students who did not suffer low back pain. They wore a cuff with a blood pressure meter on their abdomens under the pelvic belt. They held a weight for 5 s slightly above an adjustable stand at a specified forward bending angle. The external abdominal pressure (EAP) was measured 4 s after lifting the weight. The weight was 0, 10, 20 or 30 kg; the angle was 0, 30, 60 or 90 degrees; there were 4 x 4 = 16 trials/subject. As the weight increased (at all angles), EAP increased significantly. However, at 0 kg and 10 kg, EAP decreased as the angle increased; at 0 kg, EAP decreased significantly. Thus, the belt should be effective for manual material handling due to the sensitive response of EAP for low back load. With light weights such as 0 kg and 10 kg, it is better to use the belt at small bending angles than at large bending angles because EAP decreases as the angle increases.

Whereas short-term (minutes) facilitation at Aplysia sensory–motor neuron synapses is presynaptic, long-term (days) facilitation involves synaptic growth, which requires both presynaptic and postsynaptic mechanisms. How are the postsynaptic mechanisms recruited, and when does that process begin? We have been investigating the possible role of spontaneous transmitter release from the presynaptic neuron. In the previous paper, we found that spontaneous release is critical for the induction of long-term facilitation, and this process begins during an intermediate-term stage of facilitation that is the first stage to involve postsynaptic as well as presynaptic mechanisms. We now report that increased spontaneous release during the short-term stage acts as an orthograde signal to recruit postsynaptic mechanisms of intermediate-term facilitation including increased IP3, Ca 2+ , and membrane insertion and recruitment of clusters of AMPA-like receptors, which may be first steps in synaptic growth during long-term facilitation. These results suggest that the different stages of facilitation involve a cascade of pre- and postsynaptic mechanisms, which is initiated by spontaneous release and may culminate in synaptic growth.

Long-term plasticity can differ from short-term in recruiting the growth of new synaptic connections, a process that requires the participation of both the presynaptic and postsynaptic components of the synapse. How does information about synaptic plasticity spread from its site of origin to recruit the other component? The answer to this question is not known in most systems. We have investigated the possible role of spontaneous transmitter release as such a transsynaptic signal. Until recently, relatively little has been known about the functions of spontaneous release. In this paper, we report that spontaneous release is critical for the induction of a learning-related form of synaptic plasticity, long-term facilitation in Aplysia . In addition, we have found that this signaling is engaged quite early, during an intermediate-term stage that is the first stage to involve postsynaptic as well as presynaptic molecular mechanisms. In a companion paper, we show that spontaneous release from the presynaptic neuron acts as an orthograde signal to recruit the postsynaptic mechanisms of intermediate-term facilitation and initiates a cascade that can culminate in synaptic growth with additional stimulation during long-term facilitation. Spontaneous release could make a similar contribution to learning-related synaptic plasticity in mammals.

Long-term potentiation in hippocampal neurons has stages that correspond to the stages of learning and memory. Early-phase (10–30 min) potentiation is accompanied by rapid increases in clusters or puncta of presynaptic and postsynaptic proteins, which depend on actin polymerization but not on protein synthesis. We have now examined changes in pre- and postsynaptic puncta and structures during glutamate-induced late-phase (3 hr) potentiation in cultured hippocampal neurons. We find that (1) the potentiation is accompanied by long-lasting maintenance of the increases in puncta, which depends on protein synthesis, (2) most of the puncta and synaptic structures are very dynamic, continually assembling and disassembling at sites that are more stable than the puncta or structures themselves, (3) the increase in presynaptic puncta appears to be due to both rapid and more gradual increases in the number of sites where the puncta may form, and also to the stabilization of existing puncta, (4) under control conditions, puncta of postsynaptic proteins behave similarly to puncta of presynaptic proteins and share sites with them, and (5) the increase in presynaptic puncta is accompanied by a similar increase in presumably presynaptic structures, which may form at distinct as well as shared sites. The new sites could contribute to the transition between the early and late phase mechanisms of plasticity by serving as seeds for the formation and maintenance of new synapses, thus acting as local “tags” for protein synthesis-dependent synaptic growth during late-phase plasticity.

Vascular endothelial growth factor (VEGF) is implicated as a molecular mediator for adult neurogenesis and behavioral effects of antidepressant drugs. However, these potential roles of VEGF in the CNS have not been clarified in model animals. Here we have created transgenic mice overexpressing a short active variant of VEGF-A (VEGF120) in forebrain. Expression of VEGF120 significantly enhanced cell proliferation and angiogenesis, as exemplified by the formation of an enlarged reddish brain. Adult neurogenesis in hippocampus was markedly stimulated without affecting cell differentiation of neural progenitor cells. Hippocampal neurogenesis was particularly robust in young adult animals, but it declined with age and reduced to control levels by 20 weeks under continuous expression of VEGF120. Thus, VEGF alone is not sufficient to support the long-term enhancement of adult neurogenesis, and VEGF-induced vascularization per se does not necessarily predict increased neurogenesis. In transgenic mice, we observed significant changes in affective behaviors. VEGF was found to have not only antidepressant effects but also anxiolytic effects. In addition, we found that VEGF significantly reduced fear and aggression. In contrast, basal activities under natural conditions were not affected much. Unexpectedly, these characteristic behaviors were maintained in older transgenic mice undergoing a reduced level of cell proliferation in hippocampus, suggesting that there is potential dissociation between adult neurogenesis and mood regulation. Our data indicate that VEGF exerts strong neurogenic and angiogenic effects in postnatal brain and influences different forms of affective behaviors.

In this study, we evaluated the effects on subjective discomfort among cooks during food preparation through use of a standing aid that we developed to alleviate the workload on the low back in the forward-bent posture. Twelve female cooks who worked in a kitchen in a nursing home were asked to prepare foods in 2 working postures: (a) supported by the standing aid (Aid) and (b) without the aid (No aid). They were instructed to evaluate discomfort in 13-body regions during food preparation and the degree of fatigue at the day's end and to enter their ratings after the end of the workday. Since a significant correlation was observed between body height and the improvement effect of discomfort through use of the standing aid, cooks were divided into two groups according to the height, and ratings were analyzed in each group. Among the tall cooks, subjective discomfort in the low back and the front and back of thighs was significantly less with the Aid posture than with the No aid posture. However, in short cooks these values tended to increase in the Aid posture compared with the No aid posture. The results suggest that the standing aid was effective in alleviating tall cooks' workload on the low back in the forward-bent posture.

Activity-dependent re-organizations of central synapses are thought to play important roles in learning and memory. Although the precise mechanisms of how neuronal activities modify synaptic connections remain to be elucidated, the activity-induced neuronal proteins such as Homer1a may contribute to the onset of synaptic remodeling. To further understand the physiological roles of Homer1a, we first examined prolonged effects of neuronal stimulation capable of inducing Homer1a on the distribution of a postsynaptic protein Homer1c by live imaging and immunostaining. We found that glutamate stimulation induced a biphasic change in the distribution of Homer1c, in which the postsynaptic clusters of Homer1c defused initially after 30 min to 1 h, and then reassembled more than the original level after 4–8 h. When other synaptic proteins (postsynaptic density-95 (PSD95), Filamentous actin (F-actin), glutamate receptors, synaptotagmin, synaptophysin and synapsin) were analyzed by immunocytochemical methods, the distribution of these proteins also showed a similar biphasic pattern, suggesting that glutamate stimulation induces a global alteration in synaptic structures. To further dissect the functions of Homer1a in the activity-induced synaptic remodeling, the short hairpin RNA (shRNA) vectors that specifically block the expression of endogenous Homer1a were constructed. When the shRNA of Homer1a was introduced to the cells, the activity-induced changes were almost completely suppressed. The expression of surface glutamate receptor 2 was also inhibited, suggesting that Homer1a may modulate the efficacy of synaptic transmission. Furthermore, we found that Homer1a contributes to the presynaptic remodeling in a retrograde manner. Our data indicate that Homer1a regulates the activity-induced biphasic changes of post- and pre-synaptic sites.

Pkn2 is a putative transmembrane protein serine/threonine kinase required for normal development of Myxococcus xanthus. The effect of Pkn2 overexpression on development of M. xanthus was examined by expressing pkn2 under the control of a kanamycin promoter. Pkn2 was clearly detected by Western blot (immunoblot) analysis in the overexpression strain (the PKm/pkn2 strain) but could not be detected in the wild-type strain. Overexpressed Pkn2 was located almost exclusively in the membrane fraction, suggesting that Pkn2 is a transmembrane receptor-type protein Ser/Thr kinase. The PKm/pkn2 strain formed fruiting bodies more slowly than the wild-type strain, in contrast to a Pkn2 deletion strain, the delta pkn2 strain, which developed faster than the wild-type strain. However, spore production was reduced in both the PKm/pkn2 and delta pkn2 strains. These data suggest that Pkn2 functions as a negative regulator for fruiting-body formation and that the proper level of Pkn2 is necessary for maximum myxospore yield.

Gonadotropin-releasing hormone (GnRH) is the central regulator of the reproductive axis. Normal sexual maturation depends on the migration of GnRH neurons from the olfactory placode to the hypothalamus during development. Previously, we showed restricted expression of the membrane receptor adhesion-related kinase (Ark) in immortalized cell lines derived from migratory but not postmigratory GnRH neurons. In addition, Ark and GnRH transcripts were detected along the GnRH neuron migratory route in the E13 mouse cribriform plate. In the present study, we examined the role of Ark and its ligand, Gas6 (encoded by growth arrest-specific gene 6), in GnRH neuron migration. Gas6 stimulated lamellipodial extension, membrane ruffling, and chemotaxis of immortalized NLT GnRH neuronal cells via the Ark receptor. Gas6/Ark signaling promoted activation of the Rho family GTPase Rac, and adenoviral-mediated expression of dominant negative N17Rac abolished Gas6/Ark-induced actin cytoskeletal reorganization and migration of GnRH neuronal cells. In addition, p38 MAPK was activated downstream of Ark and Rac, and inhibition of p38 MAPK with either SB203580 or adenoviral dominant negative p38alpha also blocked Gas6/Ark-mediated migration. Finally, downstream of Rac and p38 mitogen-activated protein kinase (MAPK), Gas6/Ark signaling promoted activation of MAPK-activated protein kinase 2 and induced phosphorylation of HSP25, a known regulator of cortical actin remodeling. The data are the first to demonstrate a migratory signaling pathway downstream of Ark/Axl family receptors and suggest a previously unidentified role for p38 MAPK in neuronal migration. Furthermore, these studies support a potential role for Ark in the regulation of GnRH neuronal migration.

A change in the efficiency of synaptic communication between neurons is thought to underlie learning. Consistent with recent studies of such changes, we find that long-lasting potentiation of synaptic transmission between cultured hippocampal neurons is accompanied by an increase in the number of clusters of postsynaptic glutamate receptors containing the subunit GluR1. In addition, potentiation is accompanied by a rapid and long-lasting increase in the number of clusters of the presynaptic protein synaptophysin and the number of sites at which synaptophysin and GluR1 are colocalized. These results suggest that potentiation involves rapid coordinate changes in the distribution of proteins in the presynaptic neuron as well as the postsynaptic neuron.

In order to estimate occupational risk factors for health problems among truck drivers, a questionnaire survey of working conditions, job content in truck transportation, subjective symptoms and present illnesses was carried out among 541 truck transportation workers in 1997. The valid response rate was 85.7%, and 134 local truck drivers, 199 long-distance truck drivers and 71 clerical workers were analyzed. First, to examine occupational risk factors and health problems among the three groups, the authors analyzed working conditions, job content in truck transportation, subjective symptoms and present illnesses. Second, to estimate the work-relatedness of health problems among local truck drivers and long-distance truck drivers, logistic regression analyses were conducted, and odds ratios and 95% confidence intervals were computed. The prevalence rates of working factors affecting health problems of truck drivers were significantly higher than those of clerical workers in the items on irregular shift work, working environment, working posture, handling heavy materials, job stress due to overloading and long working time and limited time off. The prevalence rates for subjective symptoms (ringing in the ears, neck pain and low back pain) and present illnesses (hypertension, ulcers in the digestive tract, back injuries, whiplash injuries and hemorrhoids) among truck drivers were significantly higher than those of clerical workers. In logistic regression analyses, many work-related items except age, BMI and smoking habit showed significantly higher odds ratios for subjective symptoms and present illnesses of truck drivers. Odds ratios for hypertension, heart diseases and related subjective symptoms among local truck drivers were significantly increased by job career, twisting posture, vibration and driving stress. Odds ratios for gastro-duodenal diseases and related subjective symptoms were significantly increased by narrow working space, sleeping in the truck, driving distance, squatting posture and driving stress. Odds ratios for ringing in the ears among local truck drivers were significantly increased by job career, long working time, narrow working space, sleeping in the truck and driving stress. Odds ratios for musculo-skeletal diseases and related subjective symptoms were significantly increased by overwork, vibration, narrow working space, sitting posture and shortage of recess. Odds ratios for fatigue symptoms were significantly increased by the shortage of recess, vibration and driving stress. In order to cope with the health problems of truck drivers, it is recommended that working conditions and work loads for among truck drivers as described above be improved. Language: ja

The purpose of this study was to examine the possible effects of a tilting seat on the back, lower back and legs. Ten healthy male subjects aged 22-28 performed word-processing operations while sitting on a chair for one hour under two different seating conditions: the rocking condition and the fixed condition. While the subjects were performing the task, measurements of lower leg swelling were taken using bioelectrical impedance plethysmography, and pain scores were recorded every five min for the neck, shoulders, back, lower back, hips and legs. Electromyograms (EMGs) of the back and lower back (at Th5-6, Th8-9, L1-2 and L3-4) were recorded every sec. In addition, the subjects were videotaped while using the rocking seat, in order to analyze the angle and frequency of seat tilting. At the end of the experiment, the subjects were asked to evaluate the two conditions with respect to localized fatigue and operational efficiency. There was no significant difference in lower leg swelling between the two conditions. EMGs were significantly different at Th5-6, Th8-9 and L1-2 between the two conditions. The rocking condition generated greater EMGs at Th5-6 and L1-2, whereas the fixed condition produced greater EMGs at Th8-9. The pain scores for the neck, shoulders, back and lower back were higher under the fixed condition, while those for the buttocks were higher under the rocking condition. The average tilting frequency was as low as 25.2 times per hour, with 15.6 times per hour for tilting angles ranging from 1 to 2 degrees, and 9.6 times per hour for tilting angles exceeding 2 degrees. As for the subjective evaluations of localized fatigue, seven of the ten subjects preferred the rocking condition, while two preferred the fixed condition and one subject had no preference. Thus, there was a significant difference in the subjective evaluations of the two chairs. These findings suggest that the rocking condition, in contrast to the fixed seating condition, reduced back and lower back pain as a result of its tilting capability. The results of EMGs suggest that the rocking condition reduced back and lower back pain by increasing the overall muscle activity of the back and lower back. The leg impedance measurements showed no effect of the rocking condition on the leg swelling, as compared with the fixed condition.

A gene, pkn2, encoding a Myxococcus xanthus protein with significant similarities to eukaryotic protein serine/threonine kinases, was cloned using the polymerase chain reaction. The open reading frame for the protein, beginning with a GUG initiation codon, consists of 830 amino acids. The amino-terminal 279 residues show 37% identity to catalytic domain of Pkn1, another protein serine/threonine kinase expressed during the development at the onset of sporulation. The catalytic domain of Pkn2 contains 27% and 25% identity to rat Ca2+/calmodulin-dependent protein kinase and Bos taurus rhodopsin kinase, respectively. In the middle of the carboxy-terminal regulatory domain, there is a typical transmembrane domain consisting of 18 hydrophobic residues. The gene product, Pkn2, produced in Escherichia coli under a T7 promoter was phosphorylated at both serine and threonine residues. TEM-beta-lactamase produced in E. coli was found to serve as an effective substrate for Pkn2, phosphorylated only at threonine residues, shifting its apparent molecular mass from 29 to 44 kD. The phosphorylated beta-lactamase was unable to be secreted into the periplasmic space and localized in the cytoplasmic and membrane fractions. Analysis of phoA fusions with pkn2 demonstrated that Pkn2 is a transmembrane protein with the kinase domain in the cytoplasm and the 207-residue carboxy-terminal domain outside the cytoplasmic membrane. Disruption of pkn2 showed no effect on vegetative growth but reduced the yield of myxospores by 30%-50%. On the basis of the present results, we propose that Pkn2 is a transmembrane protein serine/threonine kinase that regulates the activity of endogenous beta-lactamase or related enzymes in response to an external signal yet to be identified.

In order to establish a measuring method of electromyogram (EMG) to evaluate the low back load by handling weight and forward bending angle. EMG at Th5-6, Th8-9, Th11-12, L1-2 and L3-4 were measured in ten male subjects. The handling weight was changed at 0, 10, 20 and 30 kg, and the forward bending angle was changed at 0, 30, 60 and 90 degree. The following results were obtained: 1) EMG increased according to the handling weight at all forward bending angles. The increasing rate of EMG at Th5-6 was larger than that at L1-2 or L3-4. Thus, EMG at higher position such as Th5-6 was found to be appropriate to evaluate handling weight-load. 2) The effect of forward bending angle was different by handling weight. EMG at L1-2 and L3-4 increased according to forward bending angle when the handling weight was 0 kg or 10 kg and when the forward bending angle was not greater than 60 degree. However, when the handling weight was 20 kg or 30 kg, EMG and forward bending angle did not show a simple proportional relationship. Thus, EMG at L1-2 or L3-4 could be used to evaluate posture load when the handling weight was light and the bending angle was not so large. However, it was difficult to use EMG to evaluate posture load when the forward bending angle was large or the handling weight was heavy.

We have developed a new armrest (NA) with adequate width supporting forearms, inclining upper surface, an area for mouse, and fixers at the edge of a shallow desk. We compared the cervicobrachial load among NA, NA with a mouse-pad (NAM) and a conventional palmrest with a mouse-pad (PRM) during word processing work. Seventeen healthy subjects operated a word processor for 1-h, alternately using 3 devices. EMG and subjective pain scores (PS) were recorded mainly from the right-side upper limb. EMG of the trapezius in PRM was significantly lower than in NAM; EMG of the anterior deltoideus (DELTa) in NA lower than in NAM; EMG of the medial deltoideus (DELTm) in NA lower than in PRM. The shoulder PS in NA/NAM was significantly lower than in PRM; the upper arm PS in NAM lower than in PRM; the forearm PS in NA/NAM lower than in PRM. These results suggest that NA reduces the cervicobrachial load than PRM.

The purpose of this study is to compare a New pen (N pen) with a conventional pen (C pen) for the load on the upper limb during 1 h writing. EMG of flexor pollicis brevis and the pain scores for thumb, forefinger, middle finger, forearm and shoulder were significantly lower for N pen than for C pen.