A new approach to spike sorting for multi-neuronal activities recorded with a tetrode--how ICA can be practical

Multi-neuronal recording with a tetrode is a powerful technique to reveal neuronal interactions in local circuits. However, it is difficult to detect precise spike timings among closely neighboring neurons because the spike waveforms of individual neurons overlap on the electrode when more than two neurons fire simultaneously. In addition, the spike waveforms of single neurons, especially in the presence of complex spikes, are often non-stationary. These problems limit the ability of ordinary spike sorting to sort multi-neuronal activities recorded using tetrodes into their single-neuron components. Though sorting with independent component analysis (ICA) can solve these problems, it has one serious limitation that the number of separated neurons must be less than the number of electrodes. Using a combination of ICA and the efficiency of ordinary spike sorting technique (k-means clustering), we developed an automatic procedure to solve the spike-overlapping and the non-stationarity problems with no limitation on the number of separated neurons. The results for the procedure applied to real multi-neuronal data demonstrated that some outliers which may be assigned to distinct clusters if ordinary spike-sorting methods were used can be identified as overlapping spikes, and that there are functional connections between a putative pyramidal neuron and its putative dendrite. These findings suggest that the combination of ICA and k-means clustering can provide insights into the precise nature of functional circuits among neurons, i.e. cell assemblies.     

Development of early apopotosis and changes in lymphocyte subsets in lymphoid organs of mice orally inoculated with nivalenol

Development of early apoptosis and changes in lymphocyte subsets were examined in lymphoid organs of female BALB/c mice after oral administration of 15 mg/kg b.w. of nivalenol (NIV), the major type B trichothecene mycotoxin, by FACS analysis. Judging from the results of viable cell count and apoptotic cell index, NIV attacked Peyer's patches first and thymus most severely. In thymus, selective damage in CD4(+)CD8(+) cells was observed at 12 and 24 h after inoculation (HAI), following the peak of apoptosis at 9 HAI. CD4(+) cells were clearly suppressed at 3 HAI in Peyer's patches, at and after 9 HAI in mesenteric lymph nodes, and 3 to 12 HAI in spleen, respectively. CD8(+) cells were also suppressed at 24 HAI in mesenteric lymph nodes and at 12 HAI in spleen, respectively. As to changes in B cell subsets, IgG(+) cells significantly decreased from 3 to 12 HAI and all B cell subsets at 24 HAI in mesenteric lymph nodes. In spleen, IgM(+) cells were suppressed at 9 HAI. On the other hand, in Peyer's patches, following clear decrease in the numbers of pan-T and pan-B cells and viable cells at 3 HAI, all B cell subsets, especially IgA(+) cells, showed a significant increase in their numbers at 9 HAI, and the numbers of IgA(+) and IgM(+) cells remained higher values than controls thereafter. Taken together, in the course of recovery from NIV-induced prominent damage in Peyer's patches at 3 HAI, interaction of NIV with Peyer's patches might result in in vivo stimulation of interleukin production at this site and result in increased proliferation and differentiation of IgA-secreting B cells at and after 9 HAI.     

Association of HLA alleles with response (especially biochemical response) to interferon therapy in Japanese patients with chronic hepatitis C.

The response of chronic hepatitis C to interferon (IFN) treatment is classified as complete response (CR), biochemical response (BR), or no response (NR). Several studies have found no difference in prevention of hepatocellular carcinoma by IFN therapy between patients with CR and those with BR. We investigated whether specific human leukocyte antigen (HLA) alleles were associated with response to IFN, especially BR, in 138 patients with chronic hepatitis C. Comparing patients with and without CR, male, a low viral titer, genotype 2a or 2b, HLA-B55, and HLA-DRB1-0803 were more common in the group with CR. Multivariate analysis showed that age (adjusted odds ratio [OR], 0.95 by every year [95% confidence interval [CI] 0.90 - 0.99], p = 0.028), genotype 2a or 2b (5.21 [95% CI 1.63 - 16.6], p = 0.005), and low viral titer (8.58 (2.66 - 27.7), p < 0.001) were associated with CR. Comparing patients with BR and NR, the pretreatment alanine aminotransferase (ALT) level was lower in the BR group (p < 0.001). Both HLA-B7 and HLA-DRB1-0101 were more common in this group (p = 0.002). As the alleles HLA-B7 and HLA-DRB1-0101 were in linkage disequilibrium, the HLA-B7-DRB1-0101 haplotype may be associated with BR. Multivariate analysis indicated that a low ALT level (0.98 by every 1 IU/L [95% CI 0.98 - 0.99], p = 0.001) and HLA-B7-DRB1-0101 haplotype (32.3 [95% CI 1.50 - 693.1], p = 0.026) contributed significantly to BR. This study suggested that host HLA expression, but not viral factors, can influence BR.     

Age-related changes in the levels of voltage-dependent calcium channels and other synaptic proteins in rat brain cortices

Neurotransmitter release from synapses is one of the most important interneuronal signaling in the nervous system. We previously reported that aging decreases depolarization-induced acetylcholine release in rat brain synaptosomes. To investigate the mechanisms underlying the age-related decrements of neurotransmission, we determined the levels of the alpha1 subunit proteins of voltage-dependent calcium channels (VDCCs) and three synaptic proteins that relate to exocytotic processes using synaptosomes prepared from cerebral cortices of young (6-month-old) and aged (27-month-old) rats. Immunoblotting analyses revealed that the protein levels of alpha1A (P/Q-type) and alpha1B (N-type) subunits in aged rats were 38% and 43% lower than the levels of young rats, respectively, but the levels of the alpha1C (L-type) subunit were not different between young and aged. On the contrary, the levels of synaptotagmin-1, synaptophysin and syntaxin were not significantly different between the two age groups in the synaptosomal preparations. These results suggest that synaptic density does not change much in the cerebral cortex in normal aging, and that the reduction of P/Q-type and N-type VDCCs, both of which participate in neurotransmitter release, is one of the causes for the decrease of neurotransmission at aged synapses.     

Numerous eosinophilic globules (skeinoid fibers) in a duodenal stromal tumor: An exceptional case showing smooth muscle differentiation

A unique case of duodenal stromal tumor In a 51-year-old man is reported. The tumor histologically showed spindle cell proliferation and numerous eosinophilic globules. Most globules were composed of tangled 45 nm thick fibrils, which were ultrastructurally Identical to 'skelnoid fibers'. The presence of glycogen granules in the tumor cells and the Immunoreactivity for α-smooth muscle actin suggested smooth muscle differentiation. Focal ultrastructural findings also supported the smooth muscle nature of this tumor. There were no immunohistochemical and ultra-structural features indicating neural differentiation. In previous studies, the presence of such 'skeinoid fibers' was suggested to be a histological marker for neural differentiation in gastrointestinal stromal tumor. However, the findings In the present case suggest that numerous 'skeinoid fibers' can be Identified in duodenal stromal tumor with smooth muscle differentiation, although this condition may be rare.     

Functional CD4+T cell subsets defined by expression of CD45RC and NTA260 antigens and age-associated polarization in murine lupus

Using two mAb, one specific to the alternative exon 6-dependentepitope of CD45 molecules(JH6.2) and one a natural thymocytotoxicautoantibody (NTA) with an unknown reactive epitope (NTA260),we subdivided splenic CD4⁺ T cells from 2-month-old BALB/c miceinto five phenotypically distinct subsets. CD45RC⁺NTA260^–(SI) cells were phenotypically analogous to CD4⁺ T cells predominatingin newborn mice and produced a significant amount of IL-2, butnot so IL-4, IL-10 or IFN- when stimulated with immobilizedanti-CD3 mAb in vitro. They appeared to consist mainly of naiveT_hP cells. The CD45RC⁺;NTA260⁺ (S II) subset also produced IL-2,but not other cytokines; however, the IL-2 levels produced weremuch higher than seen with the S I subset, thereby suggestingthe predominance of further maturated T_hP cells. The D45RC^–NTA260⁺(S III) subset mainly produced IL-4, IL-10, IFN- and less IL-2,and contained memory cells that helped the secondary antibodyresponse to a recall antigen, and hence contained T_h2 and probablya mixture of T_h0 and T_h1 cells. The CD45RC^–NTA260^–(S IV) subset was a poor responder to the immobilized anti-CD3mAb. The CD45RC^brightNTA260^dull(S V) subset consisted of a smallnumber of cells that were phenotypically analogous to activatedCD4⁺ T cells. While an age-associated decrease in the proportionof S I and less markedly in S II and in turn increase in S IIIsubsets of CD4⁺ T cells occurred in normal BALB/c mice, autoimmunedisease-prone (NZBxNZW)F₁ mice showed a marked age-associateddecrease in the proportion of not only S I, II but also IIIsubsets. As aged (NZBxNZW)F₁ mice carry CD4⁺ T helper cellsfor IgG anti-DNA antibody production, such age-associated polarizationto the S IV subset appears to be critical in the pathogeneslsof autoimmune disease in these mice.     

Skeletal manifestations in Fryns syndrome

We report on a female baby with Fryns syndrome who died soon after birth. The patient had short limbs, coarse face, hypoplastic lungs, diaphragmatic hernia, and acral hypoplasia. Literature review disclosed varying degrees of skeletal manifestations in Fryns syndrome; short limbs may be a component of Fryns syndrome. © 1995 Wiley-Liss, Inc.     

GRO-alpha in Human Serum: Differences Related to Age and Sex

PROBLEM: Human GRO-alpha (GRO-α) is a new member of the chemokine family that is supposed to play an important role in inflammatory and immune reactions. We established a sandwich enzyme-linked immunoassay (ELISA) system with polyclonal antibodies against human GRO-α and investigated the serum level of healthy donors to establish normal ranges for this chemokine in adults. METHODS: GRO-α concentrations were measured cross-sectionally in the sera of 240 healthy adults. The variability of serum GRO-α levels was also measured in normal volunteers, samples from whom were obtained by sequential venipunctures or by a small plastic cannula with a heparin-saline lock, to determine short-term variability. RESULTS: Whereas there was no difference between the concentration of human GRO-α from men (logarithmic mean, 77.6 pg/ml, n = 120) and that from women with normal menstrual cycles (log mean, 71.6 pg/ml, n = 73), the concentration from postmenopausal women (log mean 45.0 pg/ml, n = 31) was lower than that from women with normal menstrual cycles (log mean 71.6 pg/ml, n = 73). However, we could not detect any significant difference between healthy donors' serum levels and those of donors with acute inflammation. Fewer variations were recognized in the case of the sequential venipunctures method than in that of the heparin-saline lock method. CONCLUSION: We found that the GRO-α concentration of postmenopausal women was significantly lower than that of women with normal menstrual cycles. These results suggest the GRO-α serum levels of normal healthy women may have some correlation with sex hormones.