Counter-regulation of the AP-1 monomers pATF2 and Fos: Molecular readjustment of brainstem neurons in hearing and deaf adult rats after electrical intracochlear stimulation.

Expression of the immediate-early gene fos (also known as c-fos) and phosphorylation of the product of the early response gene atf2 (pATF2) in the adult auditory brainstem can be modulated by electrical intracochlear stimulation. The Fos and pATF2 proteins are competitive monomers of the heterodimeric activator protein-1 (AP-1) transcription factor that triggers the expression of genes related to neural plasticity. Our previous findings showed that the stimulation-induced spatio-temporal pattern of Fos expression in the adult auditory system depends on hearing experience. In this study, we aimed to identify a possible correlation of pATF2 and Fos expression. Adult normal hearing and neonatally deafened rats were unilaterally stimulated with a cochlear implant (CI) for 45 min, 73 min, or 2h. The numbers of Fos- and pATF2-positive neurons in the anteroventral cochlear nucleus (AVCN), the lateral superior olive (LSO), and the central inferior colliculus (CIC) were evaluated. Following stimulation, an increased Fos expression was demonstrated in all these regions in hearing and deaf rats. However, in neonatally deafened rats, significantly more Fos-positive neurons emerged that did not obey a tonotopic order. Independent of hearing experience, Fos expression correlated with a locally matching decrease of pATF2 expression in AVCN and LSO, but not in CIC. We suggest that these changes in gene expression result in a shift of AP-1 dimer composition from ATF2:Jun to Fos:Jun. This change in AP-1 constellation is expected to invoke different transcriptional cascades leading to distinct modes of tissue reorganization and plasticity responses in the mature central auditory system under stimulation.

[The utilization of brain plasticity by cochlear implants : Molecular and cellular changes due to electrical intracochlear stimulation]. / Nutzung der Plastizität des Gehirns durch Cochleaimplantate : Molekulare und zelluläre Veränderungen durch elektrische intracochleäre Stimulation.

BACKGROUND AND OBJECTIVES: During pre- and postnatal development, a high level of growth-associated protein 43 (Gap43) is expressed in the brain. This neuron-specific protein is expressed in somata, axons, and growth cones and plays a key role in neurite outgrowth and synaptogenesis. With maturation of the brain, Gap43 is down-regulated by most neurons, except in brain areas such as the hippocampal CA3 region or the binaural auditory regions lateral superior olive (LSO) and central inferior colliculus (CIC). This study investigated how changes in sensory activity levels and patterns can modulate the adult plasticity response. METHODS: To study the effect of sensory activity on adult Gap43 expression, mRNA and protein levels were determined in LSO and CIC of hearing-experienced rats, unilaterally and bilaterally deafened rats, or rats unilaterally stimulated by a cochlear implant (CI). RESULTS: Unilateral hearing loss of an adult auditory system causes asymmetrical expression of Gap43 mRNA between ipsi- and contralateral LSOs or CICs of the brain stem. While the mRNA level rose on the contralateral side of the LSO, CIC neurons increased their gap43 transcription ipsilaterally compared to the control level (p<0.001). Compensation of the lost sensory input by way of CI stimulation resulted in a bilaterally symmetric but increased gap43 transcription. CONCLUSIONS: Our data indicate that Gap43 is not only a marker for neuronal growth and synaptogenesis, but also reflects modified patterns of synaptic activities on auditory neurons. Thus, unilateral deafness directly results in an asymmetrical adaptation of the gap43 transcription between both sides of the auditory brain stem. This can be prevented by simple-patterned stimulation of the auditory nerve via a CI.

A major role for host MHC class I antigen presentation for promoting islet allograft survival.

The purpose of this study was to determine the role for CD8 T cells versus generalized MHC class I-restricted antigen presentation in islet allograft rejection and tolerance. Diabetic C57BI/6 (B6, H-2(b)) controls, C57BI/6 CD8-deficient (CD8 KO), or MHC class I-deficient C57BI/6 (beta 2m KO) recipients were grafted with allogeneic BALB/c (H-2(d)) islets. Islet allografts were acutely rejected in untreated B6, CD8 KO, and in beta 2m KO mice, indicating that neither CD8 T cells nor host MHC class I is required for allograft rejection. We then determined the efficacy of costimulation blockade in these same strains. Costimulation blockade with anti-CD154 therapy facilitated long-term islet allograft survival in both B6 and in CD8 KO recipients. However, anti-CD154 treated beta 2m KO recipients were completely refractory to anti-CD154 therapy; all treated animals acutely rejected islet allografts with or without therapy. Also, anti-NK1.1 treatment of wild-type B6 mice abrogated graft prolongation following anti-CD154 therapy. Taken together, results show a dramatic distinction between two forms of MHC class I-restricted pathways in allograft prolongation. Although anti-CD154-induced allograft survival was CD8 T-cell independent, an intact host MHC class I-restricted (beta 2m-dependent) pathway is nevertheless necessary for allograft survival. This pathway required NK1.1+ cells, implicating NK and/or NKT cells in promoting allograft prolongation in vivo.

Heat shock-induced arrests in different cell cycle phases of rat C6-glioma cells are attenuated in heat shock-primed thermotolerant cells.

The response kinetics of rat C6 glioma cells to heat shock was investigated by means of flow cytometric DNA measurements and western blot analysis of HSP levels. The results showed that the effects on cell cycle progression are dependent on the cell cycle phase at which heat shock is applied, leading to either G1 or G2/M arrest in randomly proliferating cells. When synchronous cultures were stressed during G0 they were arrested with G1 DNA content and showed prolongation of S and G2 phases after release from the block. In proliferating cells, HSC70 and HSP68 were induced during the recovery and reached maximum levels just before cells were released from the cell cycle blocks. Hyperthermic pretreatment induced thermotolerance both in asynchronous and synchronous cultures as evidenced by the reduced arrest of cell cycle progression after the second heat shock. Thermotolerance development was independent of the cell cycle phase. Pre-treated cells already had high HSP levels and did not further increase the amount of HSP after the second treatment. However, as in unprimed cells, HSP reduction coincided with the release from the cell cycle blocks. These results imply that the cell cycle machinery can be rendered thermotolerant by heat shock pretreatment and supports the assumption that HSP70 family members might be involved in thermotolerance development.

Heat shock effects on cell cycle progression.

In mammalian cells, short-term (acute) exposure to a moderate heat shock leads to a transient arrest of cells at mainly two cell cycle checkpoints, the G1/S and G2/M transitions. This is documented by the more or less synchronous resumption of cell cycle progression from these checkpoints during recovery. The reason for the accumulation of cells at these checkpoints may be found in activity thresholds of cyclin-dependent kinases (Cdks) at both transitions which are determined by (i) the amounts of the responsible cyclins, (ii) regulatory phosphorylation of the Cdks and (iii) the inhibition of Cdks by associated regulatory proteins (Ckis). All three regulatory systems may be subject to heat-shock-dependent changes, the amounts of Ckis, in particular, being increased. Cdk-dependent phosphorylation of the retinoblastoma protein and the subsequent release of active S-phase-specific transcription factors E2F/DP are considered as major heat-sensitive steps in cell cycle progression. Furthermore, high acute heat shock and long-term (chronic) heat treatment may lead to cell-type-specific forms of cell death. All types of responses to heat treatment are subject to adaptation after a 'priming' treatment, probably due to higher levels of heat shock proteins.

Nuclear translocation of stress protein Hsc70 during S phase in rat C6 glioma cells.

The expression and the nuclear translocation of the constitutive heat shock protein 70 (Hsc70) were determined during the cell cycle in synchronized rat astrocytomic C6 glioma cells. Cells were first shifted to the G0 by serum starvation. Twelve hours after a subsequent growth stimulation by transfer to 20% newborn calf serum, about 50% of the cells entered S phase. Western blot analysis with different monoclonal antibodies showed that only the constitutively expressed and moderately stress-activated Hsc70 is induced during serum stimulation. Maximal cellular Hsc70 content (170% of the control) was observed in early to mid S phase followed by a drastic decline while cells pass through G2/M (20% of the control). Hsp70, the major heat-inducible heat shock protein in C6 cells, is not detected in either asynchronously proliferating, serum-starved or in serum-stimulated C6 cells. Analysis of the nuclear and cytoplasmic protein fractions showed a significant increase of Hsc70 translocation into the nucleus during early S phase. These results indicate a role for Hsc70 but not for Hsp70 in the process of S phase entry and/or progression in C6 cells under physiological conditions.