The major components of neurofibrillary tangles (NFT) in Alzheimer's disease are bundles of paired helical filaments (PHF) which are primarily composed of highly phosphorylated tau proteins (PHF-tau).To further understand the mechanism of PHF accumulation in NFT, we examined the calpain-induced proteolysis of highly purified and primarily non-aggregated PHF and normal tau proteins with various contents of phosphate isolated from either fetal (F-tau) or adult human brain (N-tau). The extent of proteolysis was determined by decreases in tau immunoreactivity using Western-blot analysis and a panel of site-specific tau antibodies (Alz 50, Tau-2, Tau 14, Tau-I, AT8, E-11, AH-1 and PHF-1). We found that full-size polypeptides of N-tau and F-tau were similarly and rapidly proteolyzed in vitro by calpain (calpain 11, 3.3 units/mg protein) during a 10-min incubation at 3 0 T , and that their half lives (t,,J were 1.5 min and 1.8 min, respectively. Analysis of immunoblots suggests that full-length polypeptides of tau are first degraded into large fragments similar in size to that generated endogenously, then into smaller fragments. Since both endogenous and in-vim-generated tau fragments retained N-terminal epitopes, the results suggest that most of the calpain-sensitive sites may be located in the C-terminal half of the tau molecule. In contrast, PHF were extremely resistant to degradation and only a fivefold higher concentration of calpain (1 6.7 units/mg protein) induced partial proteolysis of PHF. A major calpain-generated fragment was a 45-kDa polypeptide derived from the C-terminal region of PHF-tau, which forms a core of filaments. The results suggest that the inaccessibility of potential calpain-digestion sites in the filament core could contribute to the resistance of PHF to calpain and subsequently lead to the accumulation of PHF in Alzheimer's disease. The results also suggest that hyperphosphorylation of tau may be mariginally involved in the resistance of PHF to degradation by calpain. Ultrastructural examination revealed that, in contrast to previous studies with trypsin, calpain did not alter the morphologic appearance of filaments; after incubation with calpain, the majority of PHF remained short and disperse and the number of PHF aggregated into NFT-like clusters was not significantly increased. The results suggest that the role of calpain in promoting the aggregation and clustering of filaments is limited.
Alzheimer’s disease (AD) is a neurodegenerative disease with complex pathological characteristics, whose etiology and pathogenesis are still unclear. Over the past few decades, the role of the extracellular matrix (ECM) has gained importance in neurodegenerative disease. In this review, we describe the role of the ECM in AD, focusing on the aspects of synaptic transmission, amyloid-β-plaque generation and degradation, Tau-protein production, oxidative-stress response, and inflammatory response. The function of ECM in the pathological process of AD will inform future research on the etiology and pathogenesis of AD.
CO 2 central chemoreceptors play an important role in cardiorespiratory control. They are highly sensitive to P CO 2 in a broad range. These two sensing properties seem paradoxical as none of the known pH-sensing molecules can achieve both. Here we show that cultured neuronal networks are likely to solve the sensitivity versus spectrum problem with parallel and serial processes. Studies were performed on dissociated brainstem neurons cultured on microelectrode arrays. Recordings started after a 3 week initial period of culture. A group of neurons were dose-dependently stimulated by elevated CO 2 with a linear response ranging from 20 to 70 Torr. The firing rate of some neurons increased by up to 30% in response to a 1 Torr P CO 2 change, indicating that cultured brainstem neuronal networks retain high CO 2 sensitivity in a broad range. Inhibition of Kir channels selectively suppressed neuronal responses to hypocapnia and mild hypercapnia. Blockade of TASK channels affected neuronal response to more severe hypercapnia. These were consistent with the pK a values measured for these K + channels in a heterologous expression system. The CO 2 chemosensitivity was reduced but not eliminated by blockade of presynaptic input from serotonin, substance P or glutamate neurons, indicating that both pre and postsynaptic neurons contribute to the CO 2 chemosensitivity. These results therefore strongly suggest that the physiological P CO 2 range appears to be covered by multiple sensing molecules, and that the high sensitivity may be achieved by cellular mechanisms via synaptic amplification in cultured brainstem neurons. Spontaneous breathing requires continuous feedback controls by respiratory gases. Whereas O 2 detection is carried out by peripheral chemoreceptors, CO 2 sensing mainly depends on the central chemoreceptors (CCRs) known to be located in several brainstem areas (Feldman et al. 2003;Putnam et al. 2004;Richerson, 2004;Guyenet et al. 2005b). A mystery about the CCRs is how brainstem neurons manage to detect P CO 2 as low as 1 mmHg, couple it to a 20-30% change in ventilation, and meanwhile cover a broad range of P CO 2 (Nattie, 1999;Putnam et al. 2004). Sensitivity is an inherent property of CO 2 /pH sensing molecules that is determined by the steepness of the pH-response curve. A steep response, however, confines the sensing molecule to a narrow pH range. In contrast, a sensing molecule that covers a wide spectrum of P CO 2 tends to have very low sensitivity. Indeed, none of the known CO 2 /pH-sensing molecules is capable of producing J. Su and L. Yang contributed equally to this work. a change in membrane potentials or cellular activity by 20-30% in response to 1 mmHg P CO 2 (Jiang et al. 2005). Exactly how the respiratory neuronal networks solve these seemingly paradoxical problems is unknown.Several groups of brainstem neurons are CO 2 chemosensitive, such as serotonergic neurons in the midline raphe nuclei, glutamatergic neurons in the retrotrapezoid nucleus, and catecholaminergic neurons in the locus coerule...
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