A novel action of stargazin as an enhancer of AMPA receptor activity

Stargazin (γ-2) is disrupted in the ataxic and epileptic mutant mouse, stargazer (stg). The striking defect in the stg cerebellum is the lack of functional AMPA receptors on granule cells. Recently, it has been reported that γ-2 and its related molecules are crucial for the surface expression, synaptic targeting and recycling of AMPA receptors, being termed collectively as the transmembrane AMPA receptor regulatory proteins (TARPs). However, it is still unclear whether TARPs directly modulate AMPA receptor activity. Here we report that coexpression of GluR1 (GluR1) with γ-2 using HEK293 cells and Xenopus oocytes markedly enhanced glutamate-induced currents. This effect was far beyond the increase of AMPA receptor surface expression and accompanied by increased glutamate affinity and subunit cooperativity. Other member of TARPs (γ-3, γ-4, and γ-8) also enhanced the current response through the AMPA receptors. The enhancing effect by γ-2 coexpression was further observed for homomeric GluR2 (GluR2) channels, which, when expressed alone, are known to produce only a small or negligible current response. These results suggest that γ-2 not only promotes AMPA receptor surface expression but also directly modulates AMPA receptor activity.     

Heterosynaptic expression of depolarization-induced suppression of inhibition (DSI) in rat hippocampal cultures

Depolarization-induced suppression of inhibition (DSI) is a transient suppression of the inhibitory synaptic transmission, observed in the hippocampus and the cerebellum, upon postsynaptic depolarization. Using rat hippocampal cultures, we examined whether DSI is confined to the inhibitory synapses on the depolarized neuron or, if DSI can spread to those on neighboring non-depolarized neurons. Whole-cell recordings were performed in 108 neuronal pairs with the following synaptic responses. Stimulation of one neuron evoked the inhibitory autaptic currents (IACs) recurrently in that neuron and also elicited the inhibitory postsynaptic currents (IPSCs) orthodromically in the other neuron. In 38 of 108 pairs, the postsynaptic depolarization caused transient suppression of IPSCs (homosynaptic DSI). In 11 of the 38 pairs exhibiting the homosynaptic DSI, the depolarization also induced suppression of IACs (heterosynaptic DSI). The heterosynaptic DSI, like the homosynaptic DSI, depended on depolarizing pulse duration and was blocked by a phorbol ester. These results suggest that DSI can spread to the synapses on a neighboring non-depolarized neuron in rat hippocampal cultures.     

Activation of Human Monocytes for Enhanced Production of Interleukin 8 During Transendothelial Migration in Vitro

Interleukin-8 (IL-8) is a chemokine for polymorphonuclear leukocytes (PMNs) and lymphocytes, which promotes the extravasation of these inflammatory cells. In this study, we investigated IL-8 synthesis induced by the adhesive interaction between monocytes and endothelial cells during transmigration and the capacity of transmigrated monocytes to produce IL-8. Cocultured human monocytes and human umbilical vein endothelial cell (HUVEC) monolayers induced the synefgistic production of IL-8, compared with cultures of either monocytes or HUVEC monolayers alone. Coculture-induced IL-8 production almost doubled after HUVECs were stimulated with IL-1. The induced IL-8 mRNA expression was consistent with the protein data, indicating the de novo synthesis of IL-8 by the coculture. Monoclonal antibodies (mAbs) against IL-8 inhibited the transendothelial chemotactic activity of the supernatants for PMNs by 55%. Immunohistochemistry revealed that both adherent and transmigrated monocytes and unstimulated HUVECs expressed IL-8 protein, whereas nonadherent monocytes did little. Transmigrated monocytes spontaneously secreted a 3.8-fold greater amount of IL-8 than the initial monocytes. Coculture-induced IL-8 production was inhibited about 30% by polyclonal Abs against IL-, IL-1, or tumor necrosis factor , while it was not affected by mAbs against intercellular adhesion molecule 1 or vascular cell adhesion molecule 1. The results suggested that adhesive interaction during the transmigration of monocytes through HUVEC monolayers activates both cell types to produce IL-8 and that transmigrated monocytes are capable of producing ample IL-8.     

Nature of optokinetic response and zonal organization of climbing fiber afferents in the vestibulocerebellum of the pigmented rabbit

Summary In pigmented rabbits anesthetized with N₂O (70%) and halothane (2–4%), Purkinje cells were extracellularly recorded in the nodulus. Large field (60°×60°) optokinetic stimulation (OKS) with constant velocity was delivered to either the ipsi- or contralateral eye, and the direction and velocity selectivities of complex spike responses were examined. To ipsilateral OKS (n = 181), the preferred direction was forward (F, n = 72), upward (U, n = 38) or downward (D, n = 10), while the remaining cells (n = 61) showed no response (N). To contralateral OKS (n = 117), the preferred direction was backward (B, n = 22), upward (U, n = 7) or downward (D, n = 22), while the rest (n = 66) showed N. Cells tested with both eyes (n = 95) fell into 8 categories based on the preferred direction to ipsi- and contralateral OKS: (1) ipsi-F and contra-B (F/B type, n = 20), (2) ipsi-F but contra-N (F/N type, n = 12), (3) ipsi-U and contra-D (U/D type, n = 15), (4) ipsi-U but contra-N (U/N type, n = 13), (5) ipsi-N but contra-D (N/D type, n = 1), (6) ipsi-D but contra-N (D/N type, n = 5), (7) ipsi-N but contra-U (N/U type, n = 6), and (8) N to both eyes (N/N type, n = 23). The optimum velocity was in the range 0.1–2.0°/s for all cells responsive to OKS. In the ventral lamella, four medio-laterally aligned zones were demonstrated. In the most medial zone (0–0.5 mm from the midline), the majority of cells showed ipsi-N or contra-N responses. In the second zone (0.5–1.5 mm), most cells preferred ipsi-F or contra-B directions. In the third zone (1.5–2.5 mm), most cells preferred ipsi-U or contra-D directions. In the most lateral zone (2.5–3.5 mm), most cells preferred ipsi-F or contra-B directions. In the dorsal lamella, a longitudinal zone characterized with cells preferring ipsi-U or contra-D directions was found about 1.5–2.5 mm from the midline. This zone seemed to be the continuation of the third zone in the ventral lamella. Cells preferring ipsi-D or contra-U directions were scattered in the medial half of both the dorsal and ventral lamellae. Most cells responsive to electrical stimulation of the contralateral optic tract (OT) preferred the ipsi-F direction and were localized in the second and the most lateral zones of the ventral lamella. As for cells activated by a climbing fiber with a branching axon to the flocculus, no characteristic feature was found in terms of the preferred direction to OKS, except that none of the cells preferring ipsi-D or contra-U directions were activated by such branching climbing fibers. The results indicate that the nodulus consists of at least four functionally distinct zones in terms of direction selectivity of visual climbing fiber afferents.     

Nature of optokinetic response and zonal organization of climbing fiber afferents in the vestibulocerebellum of the pigmented rabbit

In pigmented rabbits anesthetized with N2O (70%) and halothane (2-4%), Purkinje cells were extracellularly recorded in the flocculus. A large central visual field (60 degrees x 60 degrees) was used to optokinetically stimulate either the ipsi- or contralateral eye, and the direction and velocity selectivities of complex spike responses were examined. For optokinetic stimulation (OKS) delivered to the ipsilateral eye (n = 129), the preferred direction was forward (F, n = 57) or upward (U, n = 37), while the remaining cells (n = 35) showed no response (N). For OKS delivered to the contralateral eye (n = 107), the preferred direction was backward (B, n = 11), downward (D, n = 42) or upward (U, n = 2), and the rest (n = 52) showed N. Cells tested with both eyes (n = 89) fell into five categories based on the preferred direction to ipsi- and contralateral OKS: (1) ipsi-F and contra-B (F/B type, n = 9), (2) ipsi-F but contra-N (F/N type, n = 28), (3) ipsi-U and contra-D (U/D type, n = 13), (4) ipsi-U but contra-N (U/N type, n = 17), and (5) ipsi-N but contra-D (N/D type, n = 22). The optimum velocity was within 0.1-2.0 degrees/s for all cells. On the average, the best response was obtained at 0.2-0.5 degrees/s stimulation. All ipsi-F cells responded to electrical stimulation of the optic tract (OT), while most cells preferring ipsi-U, contra-B and contra-D directions did not respond. No characteristic feature was found in cells innervated with collateralized climbing fiber branches to the nodulus. In the flocculus, cells preferring horizontal orientation (H cells, preferring ipsi-F and/or contra-B directions) were localized in a narrow dorsoventral zone (less than 1.0 mm) along the caudal border of the rostral one third, while those preferring the vertical orientation (V cells, preferring ipsi-U and/or contra-D directions) were in two distinct narrow zones located rostral and caudal to the H cell zone. H and V cells were intermingled in the central portion of the ventral flocculus. These four zones are in good agreement with previously defined H, anterior V, posterior V and R zones, respectively. The results indicate that the subdivision of the flocculus which controls horizontal (vertical) eye movements receives information regarding movements of the visual surround in the horizontal (vertical) orientation through visual climbing fiber afferents, thus being organized in olivo-cortico-nuclear functional units for control of eye movements.     

T cell can recognize the allospecificities formed by the substitution of amino acids associated with HLA-Bw4/Bw6 public epitopes.

Our previous studies clearly showed that HLA-B35 and HLA-Bw53 differed only by the amino acids associated with HLA-Bw4/Bw6 epitopes, in that the former possessed Bw6 and the latter Bw4 epitope. It remains to be known whether T cell can discriminate HLA-B35 from HLA-Bw53, although the difference between these HLA antigens is discriminated by monospecific human alloantisera. To investigate allorecognition of these HLA antigens by T cells, anti-HLA-Bw53 cytotoxic T lymphocytes (CTLs) were generated. Anti-HLA-Bw53 cytotoxic T lymphocytes (CTLs) were generated. Anti-HLA-Bw53 bulk CTLs from an individual with HLA-B35 clearly discriminated HLA-Bw53 from HLA-B35. On the other hand, anti-HLA-Bw53 bulk CTLs from an individual without HLA-B35 revealed weak cross-reactivity with HLA-B35 and HLA-B51. The additional studies using HLA-Bw53 or HLA-B35-specific CTL clones showed that some but not all of the CTL clones definitively distinguish the difference between HLA-Bw53 and HLA-B35. Thus, the allospecificities formed by HLA-Bw4/Bw6 epitopes were discriminated by allogeneic T cells. The present study demonstrated that HLA-Bw4/Bw6 public epitopes play an important role in allorecognition of T cells.     

Conservation of genes encoding HLA-B5 and B35 cross-reactive group antigens in various races

HLA-B5 and B35 CREG antigens include HLA-B35, B51, B52, B53, and B78. Recent studies suggest that the genes encoding the HLA-B5, B35 CREG, and HLA-B58 antigens share a common ancestor. We sequenced the exons of the genes encoding HLA-B51, B53, and B58 from American black individuals and the gene HLA-B52 encoding from an Arabic individual, and compared them with previously reported sequences of HLA-B51 (B^*5101) and HLA-B52 (B^*5201) from Japanese, HLA-B53 (B^*5301) from an Algerian, and HLA-B58 (B^*5801) from a Sardinian. The sequences of the genes from the American black and Arabic individuals were identical to those from the other races. These findings support the hypothesis that these antigens have evolved prior to divergence of the major ethnic groups.     

Heat and water vapour transfer of protective clothing systems in a cold environment,measured with a newly developed sweating thermal manikin

A moveable sweating thermal manikin has recently been developed. Thermal and water-vapour resistances of three kinds of cold-protective clothing ensembles, laminated with polytetrafluoroethylene, polyurethane and without a laminate were measured, with the aid of the manikin in a cold environment of 5°C with a relative humidity of 70% and an air velocity of around 1.5 m s^–1. Two sweating rates of 65 and 130 g m^–2 h^–1 were employed. Supplied heat fluxes in both of the sweat rates ranged from 350 W m^–2 to 400 W m^–2. To maintain a comfortable condition, the skin wettedness (w) (mean weighted value) had to be kept at 0.6. The measurements obtained from the manikin when testing the three ensembles were w=0.3 (approximately) for the low sweat rate and w0.6 for the high sweat rate, irrespective of the property differences among the ensembles. In addition, the condensation in the ensembles in comparison with those calculated from an analytical equation is discussed. Condensation mass fluxes in the ensembles obtained byexperiment and those from the calculation agreed sufficiently well. Thus, distribution of the condensation in the ensembles was estimated using the equation.