摘要:

Abstract:Most of the receptors for soluble factors functioning in the hematopoietic system belong to the class I cytokine receptor family. These receptors often share common signal transducing receptor components in the same family, which explains the functional redundancy of cytokines. One typical example is a group of receptor systems for interleukin‐6 (IL‐6) and related cytokines that share gp130 as a signal transducer. This subset of cytokines, i.e., IL‐6, IL‐11, leukemia inhibitory factor, oncostatin M, ciliary neurotrophic factor, and cardiotrophin‐1, are all pleiotropic, exhibiting overlapping biological activities, and are known to function also in the neuronal system. In their receptor complexes, gp130 and ligand‐specific chains possess no intrinsic tyrosine kinase domain but are associated with members of the Jak family of cytoplasmic tyrosine kinases. The Jak kinases become activated after ligand‐induced homo‐ or heterodimerization of gp130. This activation appears to link the cell surface receptors to the nuclear genes through a series of biochemical changes, including tyrosine phosphorylation and activation of a latent cytoplasmic transcription factor called signal transducer and activator of transcri

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010001.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:Previous studies demonstrated that 9 kb of the rat tyrosine hydroxylase (TH) 5′ flanking sequence directed appropriate spatiotemporal expression of alacZreporter gene to catecholaminergic cells in the CNS of transgenic mice. In the present study, specificity of transgene expression was further extended to demonstrate cell type‐specific functional regulation oflacZexpression using manipulations known to alter endogenous TH expression. Alterations inlacZreporter expression should parallel changes in endogenous TH levels if the DNA elements mediating these functional changes of TH expression in vivo reside within the 9 kb of the TH promoter region. Naris closure induced an activity‐dependent decrease of TH expression in dopaminergic periglomerular cells in the olfactory bulb that was paralleled by down‐regulation oflacZexpression in the transgenic mice. Densitometry and image analysis were used to quantifylacZexpression following acute reserpine administration (5 mg/kg, s.c.), which up‐regulates endogenous TH. At 48 h postinjection, analysis of OD values indicated a significant increase of X‐gal staining in the locus coeruleus and ventral tegmental area but not in the substantia nigra or olfactory bulb of reserpine‐treated transgenic animals. These data showed that the 9‐kb sequence also mediates cell type‐specific transsynaptic regulation of reporter gene expression. Analysis of this transgenic animal offers a useful model system to study in vivo regulation of T

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010011.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:Alternative splicing of human tyrosine hydroxylase (TH) pre‐mRNA produces four mRNAs leading to four different TH isoforms and is thought to have important regulatory functions. We show that the diversity of TH mRNAs is greater than previously described in the autonomous nervous system: New splice junctions corresponding to the skipping of exon 3 were identified by amplification of cDNA synthesized from pheochromocytoma RNA. In all cases the reading frame was maintained. These species were assayed by RNase protection experiments; their abundance (4–6%) was comparable to that of the previously identified human TH‐3 and ‐4 species in normal adrenal medulla. However, higher levels (11–34%) of these species were found in adrenal medullas of patients suffering from progressive supranuclear palsy. Whether such changes are specific to the disease or the consequences of the stress associated with this severe neurodegeneration remains to be es

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010019.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:Theciselements mediating activation of the tyrosine hydroxylase gene by angiotensin II were examined by transfecting tyrosine hydroxylase promoter‐luciferase constructs into cultured bovine adrenal medullary cells. Angiotensin II‐responsive elements are located within −54/+25‐bp and −269/−55‐bp promoter regions and were identified, respectively, as cyclic AMP (CRE)‐ and 12‐O‐tetradecanoylphorbol 13‐acetate responsive element (TRE)‐like sequences. Unlike CRE, TRE also supports basal promoter activity. Mutations of TRE or CRE that reduced angiotensin II stimulation abolished in vitro binding of nuclear proteins to those elements, suggesting that proteins forming CRE‐ and TRE‐inducible complexes may mediate angiotensin II stimulation. The TRE is adjacent to a dyad symmetry element. Those two sites form a common regulatory unit in which the dyad symmetry element acts as a repressor of the TRE site. Isolated dyad symmetry element did not bind nuclear proteins in vitro. In supercoiled DNA it exhibited S1 nuclease sensitivity and was recognized by a DNA cruciform‐specific antibody consistent with the extrusion of a cruciform structure that overlaps with the TRE. A mutation that abolished formation of the cruciform correlated with a loss of repressor activity. We propose a novel model of tyrosine hydroxylase gene regulation in which functions of the TRE are modulated via structural t

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010026.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:Messages for subunits of voltage‐gated calcium channels were examined in the cochlea of the CBAJmouse by PCR analysis. Total RNA was extracted from the auditory organs of 16–18‐day‐old animals. After reverse transcription, resulting cDNA was amplified by PCR with primers targeted to nucleotide sequences corresponding to 12 different calcium channel subunits. PCR products representing subunit gene expression were strongly and consistently amplified for α1C, α1D, α1E, α2δ, β1, β3, and β4but not for α1A, α1B, α1S, β2, or γ. The chosen primers amplified cochlear cDNA to yield an overall pattern of bands different from that of any tissue studied thus far, in particular with respect to the α2δ and β1subunits; the α2δ product was found to be significantly shorter than the corresponding brain and skeletal muscle isoforms. Nucleotide sequencing confirmed the identity of mouse cochlear subunit cDNAs. The results suggest that L‐type and presumptive R‐type calcium channels are expressed in the mammalian cochlea and that the α2δ subunits may be coded by a cha

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010037.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:We report here the positional cloning and molecular characterization of theunc‐24gene ofCaenorhabditis elegans. This gene is required for normal locomotion and interacts with genes that affect the worm's response to volatile anesthetics. The predicted gene product contains a domain similar to part of two ion channel regulators (the erythrocyte integral membrane protein stomatin and theC. elegansneuronal protein MEC‐2) juxtaposed to a domain similar to nonspecific lipid transfer protein (nsLTP; also called sterol carrier protein 2). Sequence analysis suggests that the nsLTP‐like domain of UNC‐24 provides lipid carrier function and is tethered to the plasma membrane by the stomatin‐like domain, which may be regulatory. We postulate that UNC‐24 may be involved in lipid transfer between closely appose

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010046.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:The metabotropic glutamate receptor (mGluR) subtype 1 exists as at least three variants (−1a, −1b, and −1c) generated by alternative splicing at the C‐terminal domain. Fluorometric Ca2+measurements were used to compare the concentration dependency of agonist‐induced rises in intracellular free Ca2+concentration ([Ca2+]i) in human embryonic HEK 293 cells transiently expressing rat mGluR1a, mGluR1b, or mGluR1c. The rank order of agonist potencies was quisqualate ≫ (2S,1′S,2′S)‐2‐(carboxycyclopropyl)glycine (L‐CCG‐I)>(1S,3R)‐1‐aminocyclopentane‐1,3‐dicarboxylic acid [(1S,3R)‐ACPD] and did not differ among the splice variants. However, agonists were consistently more potent at mGluR1a than at mGluR1c and mGluR1b. In the same system, we characterized the agonist pharmacology of two chimeric rat mGluR3/1 receptors where the first and/or the second intracellular loop(s) and the C‐terminal domain were exchanged with the corresponding mGluR1a or mGluR1c sequences and that were previously shown to mediate elevations in [Ca2+]iin response to agonists. The potency of agonists was higher at the chimera having the C‐terminus of mGluR1a as compared with those having the mGluR1c C‐terminus. Both chimeric mGluR3/1 receptors had the same rank order of agonist potencies: L‐CCG‐I ≫ (1S,3R)‐ACPD ∼ quisqualate. These data support the hypothesis that the C‐terminal domain of mGluRs plays a role in determining the potency of agonis

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010058.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:The observation that delayed death of CA1 neurons after global ischemia is inhibited by protein synthesis inhibitors suggests that the delayed death of these neurons is an active process that requires new gene expression. Delayed death in CA1 has some of the characteristics of apoptotic death; however, candidate proapoptotic proteins have not been identified in the CA1 after ischemia. We studied the expression of Bax protein and mRNA, a member of thebcl‐2family that is an effector of apoptotic cell death, after global ischemia in the four‐vessel global ischemia model in the rat and compared these results with the expression of the antiapoptotic genebcl‐2.BaxmRNA and protein are both expressed in CA1 before delayed death, whereas bcl‐2 protein is not expressed. Bcl‐2 protein expression, but not that of Bax, is increased in CA3, a region that is ischemic but less susceptible to ischemic injury. In the dentate gyrus, both Bax and bcl‐2 proteins are expressed. The selective expression of Bax in CA1 supports the hypothesis that Bax could contribute to delayed neuronal death in these vulnerable neurons by an independent mechanism or by forming heterodimers with gene family members other

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010064.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:To determine whether protein kinase C (PKC) mediates release of peptides from sensory neurons, we examined the effects of altering PKC activity on resting and evoked release of substance P (SP) and calcitonin gene‐related peptide (CGRP). Exposing rat sensory neurons in culture to 10 or 50 nMphorbol 12,13‐dibutyrate (PDBu) significantly increased SP and CGRP release at least 10‐fold above resting levels, whereas the inactive 4α‐PDBu analogue at 100 nMhad no effect on release. Furthermore, 100 nMbradykinin increased peptide release approximately fivefold. Down‐regulation of PKC significantly attenuated the release of peptides evoked by either PDBu or bradykinin. PDBu at 1 nMor 1‐oleoyl‐2‐acetyl‐sn‐glycerol at 50 µMdid not alter resting release of peptides, but augmented potassium‐ and capsaicin‐stimulated release of both SP and CGRP approximately twofold. This sensitizing action of PKC activators on peptide release was significantly reduced by PKC down‐regulation or by pretreating cultures with 10 nMstaurosporine. These results establish that activation of PKC is important in the regulation of peptide release from sensory neurons. The PKC‐induced enhancement of peptide release may be a mechanism underlying the neuronal sensitizati

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010072.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Abstract:There is debate about the mechanisms mediating adenosine release from neurons. In this study, the release of adenosine evoked by depolarizing cultured cerebellar granule neurons with 50 mMK+was inhibited by 49 ± 7% in Ca2+‐free medium. The remaining release was blocked by dipyridamole (IC50= 6.4 × 10−8M) and nitrobenzylthioinosine (IC50= 3.6 × 10−8M), inhibitors of adenosine uptake. Ca2+‐dependent release was reduced by 78 ± 9% following a 21‐h pretreatment of the cells with pertussis toxin, which ADP‐ribosylates Gi/GoG proteins, thereby preventing their dissociation. The nucleoside transporter‐mediated component of K+‐induced adenosine release also was inhibited by 62 ± 8% by pertussis toxin and was potentiated by 78 ± 11% following cholera toxin treatment, which permanently activates Gs. Uptake of [3H]adenosine into cultured cerebellar granule neurons over a 10‐min period was not dependent on extracellular Na+but was reduced by dipyridamole (IC50= 3.2 × 10−8M) and nitrobenzylthioinosine (IC50= 2.6 × 10−8M). Thus, adenosine uptake likely occurs via the same transporter mediating Ca2+‐independent adenosine release. Adenosine uptake was potentiated by cholera toxin pretreatment (152 ± 15% of control), but pertussis toxin had no statistically significant effect. It is possible that Gs, Gi/Go, or free Gβγ dimer modulate the equilibrative, inhibitor‐sensitive nucleoside ca

ISSN:0022-3042    

DOI:10.1046/j.1471-4159.1996.67010081.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY