Molecular and phenotypic variation in the achaete-scute region of Drosophila melanogaster

Variation in quantitative characters underlies much adaptive evolution and provides the basis for selective improvement of domestic species, yet the genetic nature of quantitative variation is poorly understood. Many loci affecting quantitative traits have been identified by the segregation of mutant alleles with major qualitative effects. These alleles may represent an extreme of a continuum of allelic effects, and most quantitative variation could result from the segregation of alleles with subtle effects at loci identified by alleles with major effects. The achaete-scute complex in Drosophila melanogaster plays a central part in bristle development and has been characterized at the molecular level. The hypothesis that naturally occurring quantitative variation in bristle number could be associated with wild-type alleles of achaete-scute was tested by correlating phenotypic variation in bristle number with molecular variation in restriction maps in this region among chromosomes extracted from natural populations. DNA insertion variation in the achaete-scute region was found to be strongly associated with variation in bristle number.     

A novel metalloproteinase gene specifically expressed in stromal cells of breast carcinomas.

A gene has been identified that is expressed specifically in stromal cells surrounding invasive breast carcinomas. On the basis of its sequence, the product of this gene, named stromelysin-3, is a new member of the family of metalloproteinase enzymes which degrade the extracellular matrix. The suggestion is that stromelysin-3 is one of the stroma-derived factors that have long been postulated to play an important part in progression of epithelial malignancies.     

Control of neuronal fate by the Drosophila segmentation gene even-skipped

The central nervous system (CNS) contains a remarkable diversity of cell types. The molecular basis for generating this neuronal diversity is poorly understood. Much is known, however, about the regulatory genes which control segmentation and segment identity during early Drosophila embryogenesis. Interestingly, most of the segmentation and homoeotic genes in Drosophila, as well as many of their vertebrate homologues, are expressed during the development of the nervous system (for example, ref. 3). Are these genes involved in specifying the identity of individual neurons during neurogenesis, just as they specify the identity of cells during segmentation? We previously described the CNS expression of the segmentation gene fushi tarazu (ftz) and showed that ftz CNS expression is involved in the determination of an identified neuron. Here we show that another segmentation gene, even-skipped (eve), is expressed in a different but overlapping subset of neurons. Temperature-sensitive inactivation of the eve protein during neurogenesis alters the fate of two of these neurons. Our results indicate that the nuclear protein products of the eve and ftz segmentation genes are components of the mechanism controlling cell fate during neuronal development.     

Regulation of the G1-S transition in postembryonic neuronal precursors by axon ingrowth.

In the newly cellularized Drosophila embryo, progress through the cell cycle is regulated at the G2-M transition. We have examined cell-cycle regulation later in Drosophila development, in a group of postembryonic neuronal precursors. The S-phase precursor cells, which generate photoreceptor target neurons (lamina neurons) in the central nervous system, are not present in the absence of photoreceptor innervation. Here we report that axons selectively approach G1-phase precursors. Without axon ingrowth, lamina precursors do not enter their final S phase and by several criteria, arrest in the preceding G1 phase. These findings provide evidence that at this stage in development the control of cell division can occur at the G1-S transition.     

Two chemosensory receptors together mediate carbon dioxide detection in Drosophila

Blood-feeding insects, including the malaria mosquito Anopheles gambiae, use highly specialized and sensitive olfactory systems to locate their hosts. This is accomplished by detecting and following plumes of volatile host emissions, which include carbon dioxide (CO2). CO2 is sensed by a population of olfactory sensory neurons in the maxillary palps of mosquitoes and in the antennae of the more genetically tractable fruitfly, Drosophila melanogaster. The molecular identity of the chemosensory CO2 receptor, however, remains unknown. Here we report that CO2-responsive neurons in Drosophila co-express a pair of chemosensory receptors, Gr21a and Gr63a, at both larval and adult life stages. We identify mosquito homologues of Gr21a and Gr63a, GPRGR22 and GPRGR24, and show that these are co-expressed in A. gambiae maxillary palps. We show that Gr21a and Gr63a together are sufficient for olfactory CO2-chemosensation in Drosophila. Ectopic expression of Gr21a and Gr63a together confers CO2 sensitivity on CO2-insensitive olfactory neurons, but neither gustatory receptor alone has this function. Mutant flies lacking Gr63a lose both electrophysiological and behavioural responses to CO2. Knowledge of the molecular identity of the insect olfactory CO2 receptors may spur the development of novel mosquito control strategies designed to take advantage of this unique and critical olfactory pathway. This in turn could bolster the worldwide fight against malaria and other insect-borne diseases.     

Use of suppression subtractive hybridization strategy for cloning and identifying specifically expressed genes of renal cell carcinoma

Using suppression subtractive hybridization, a renal cell carcinoma (RCC) cDNA subtractive library which only contains differently expressed cDNAs between human RCC and normal kidney has been constructed. 200 clones were picked out randomly to perform enzyme digest analysis, a part of them underwent sequence analysis and Northern blot to identify RCC specially expressed genes. Results showed that 190 clones contain 50—400 bp inserts respectively. Sequence analysis was performed in 10 clones. All the 10 sequences were unknown before and derived from 6 unique novel genes among which the cDNA insert RCC18 has five copies. Northern blot analysis showed that RCC18 cDNA expressed highly in RCC, but there was no signal detected in normal kidney, and the full length of RCC18 was about 2.5 kb. The constructed cDNA subtractive library of human RCC is a highly efficient one and lays the solid foundation for large-scale screening and cloning new and specific oncogenes or tumor suppressor genes of RCC. The novel specially expressed genes provided an important clue for researching the mechanism of the occurrence and development of RCC.     

Two polyphosphatidylinositide metabolites control two K+ currents in a neuronal cell

Hydrolysis of the membrane phospholipid phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P2) produces two prospective intracellular messengers: inositol 1,4,5-trisphosphate (InsP3), which releases Ca2+ from intracellular stores; and diacylglycerol (DG), which activates protein kinase C. Here we show how the formation of these two substances triggered by one external messenger, bradykinin, leads to the appearance of two different sequential membrane conductance changes in the neurone-like NG108-15 neuroblastoma-glioma hybrid cell line. In these cells bradykinin rapidly hydrolyses PtdIns(4,5)P2 to InsP3 and DG, raises intracellular Ca2+ and hyperpolarizes then depolarizes the cell membrane. By voltage-clamp recording we show that the hyperpolarization results from the activation pharmacologically-identifiable species of Ca2+-dependent K+ current. This is also activated by intracellular injections of Ca2+ or InsP3 so may be attributed to the formation and action of InsP3. The subsequent depolarization results primarily from the inhibition of a different, voltage-dependent K+ current, the M-current that is also inhibited by DG activators. Hence we describe for the first time a dual, time-dependent role for these two intracellular messengers in the control of neuronal signalling by a peptide.     

Excitatory effect of histamine on neuronal activity of rat cerebellar fastigial nucleus in vitro

The cerebellar fastigial nucleus (FN) holds an important role in motor control and body balance. Previous studies have revealed that the nucleus is innervated by direct hypothalamocerebellar histaminergic fibers. However, the functional role of histaminergic projection in cerebellar FN has never been established. In this study, we investigated the effect of histamine on neuronal firing of cerebellar FN by using slice preparations. Sixty-five FN cells were recorded from 47 cerebellar slices, and a vast majority of the cells responded to histamine stimulation with an excitatory response (58/65, 89.2%). Perfusing slices with low-Ca2 /high-Mg2 medium did not block the histamine-induced excitation (n=10), supporting a direct postsynaptic action of histamine on the cells. Furthermore, the excitatory effect of histamine on FN neurons was not blocked by selective histamine H1 receptor antagonist triprolidine (n=15) or chlorpheniramine (n=10), but was effectively suppressed by ranitidine (n=15), a highly selective histamine H2 receptor antagonist. On the other hand, highly selective histamine H2 receptor agonist dimaprit (n=20) instead of histamine H1 receptor agonist 2-pyridylethylamine (n=16) mimicked the ex- citatory effect of histamine on FN neurons. The dimaprit-induced FN neuronal excitation was effectively antagonized by selective histamine H2 receptor antagonist ranitidine (n=13) but not influenced by se- lective histamine H1 receptor antagonist triprolidine (n=15). These results demonstrate that histamine excites cerebellar FN cells via the histamine H2 receptor mechanism and suggest that the hypotha- lamocerebellar histaminergic fibers may modulate cerebellar FN-mediated sensorimotor integration through their excitatory innervations on FN neurons.     

Formation of target-specific neuronal projections in organotypic slice cultures from rat visual cortex.

A characteristic feature of the mammalian cortex is that projection neurons located in distinct cortical layers send their axons to different targets. In visual cortex, cells in layers 2 and 3 project to other cortical areas, whereas cells in layers 5 and 6 project to subcortical targets such as the lateral geniculate nucleus. The proper development of these projections is crucial for correct functioning of the visual system. Here we show that specific connections are established in an organotypic culture system in which rat visual cortex slices are co-cultured with another slice of the visual cortex or with a thalamic slice. The laminar origin and cellular morphology in vitro of cortical projections to other cortical regions or to subcortical targets are remarkably similar to those seen in vivo. In addition, axons of projecting cells are not restricted to particular pathways, but appear instead to grow directly towards their appropriate target. These observations raise the possibility that chemotropic attraction from the target areas may play an important part in the development of the cortical projection pattern.     

Functional neuronal replacement by grafted striatal neurones in the ibotenic acid-lesioned rat striatum

In rats, striatal neuronal destruction by so-called excitotoxic amino acids, kainic acid or ibotenic acid (IA) produce neuropathological and neurochemical changes in the basal ganglia which resemble those seen in patients with Huntington's chorea. Such lesioned animals show a behavioural syndrome which is reminiscent of the cardinal symptoms of the disease, accompanied by a substantial increase in local cerebral metabolic activity in several striatal target structures within the extrapyramidal motor system. The study was designed to explore the potential of grafted fetal striatal neurones implanted into the IA-lesioned striatum to compensate for the structural, neurochemical, metabolic and behavioural defects of IA-lesioned rats. Extending previous studies, we report here that such striatal implants can significantly ameliorate the lesion-induced locomotor hyperactivity and at least partly normalize the metabolic hyperactivity in the extrapyramidal neuronal system.     

Achyranthes bidentata Blume extract promotes neuronal growth in cultured embryonic rat hippocampal neurons

We have prepared an aqueous extract of Achyranthes bidentata Blume, a commonly prescribed Chinese medicinal herb, and reported, in previous studies, that A. bidentata extract benefits nerve growth and prevents neuron apoptosis. In this study, we investigated the actions of A. bidentata extract on survival and growth of primarily cultured rat hippocampal neurons. The morphological observation revealed that neurite growth from hippocampal neurons was significantly enhanced by A. bidentata extract with similar effects to those induced by nerve growth factor (NGF), and the greatest neurite growth appeared on treatment with A. bidentata extract at 1 μg/ml for 24 h. DNA microarray analysis indicated that there were 25 upregulated genes and 47 downregulated genes exhibiting significantly differential expression in hippocampal neurons treated with A. bidentata extract at 1 μg/ml for 6 h when compared to those in untreated hippocampal neurons. Real-time quantitative RT-PCR and Western blot analysis demonstrated that the expression of growth-associated protein-43 in hippocampal neurons was upregulated at both mRNA and protein levels after treatment with A. bidentata extract, and the optimal dosage of the extract was also 1 μg/ml. These data confirm that A. bidentata extract could promote in vitro hippocampal neuronal growth in a dose- and time-dependent manner.     

Modulation of neuronal activity of cerebellar fastigial nucleus by locus coeruleus stimulation in the rat

The effects of stimulating locus coeruleus (LC) on neuronal activity of cerebellar fastigial nucleus (FN) was investigated. Stimulation of LC elicited inhibitory, excitatory and biphasic (inhibition-excitation) responses from FN cells. The majority of responsive cells showed an inhibitory response with a latency of less than 10 ms. Injection of α adrenoreceptor antagonists phentolamine (ⅳ) could block the inhibitory response of FN cells to the LC stimulation, but propranolol (ⅳ), a β adrenoreceptor antagonist, could not. These results suggest that LC-cerebellar noradrenergic afferent fibers may be involved in the cerebellar sensorimotor integration process by exerting their modulatory action on the cerebellar nuclear cells' activities.     

果蝇DNA和基因间序列的两个信息参数沿染色体分布

以果蝇染色体的全序列和基因间序列作为研究对象.分别用描述序列构成的两个信息参数X和F分析两类序列沿着染色体分布特征,并着重讨论染色体着丝粒及其周围区域的分布规律,用线性拟合和多项式拟合分析染色体各类序列沿染色体的统计分布特征(学生t检验),分析各个参数的平均值在染色体不同区域的分布.     

Excitatory effect of histamine on neuronal activity of rat cerebellar fastigial nucleus Jn vitro

The cerebellar fastigial nucleus （FN） holds an important role in motor control and body balance. Previous studies have revealed that the nucleus is innervated by direct hypothalamocerebellar hletaminergic fibers. However, the functional role of histaminergic projection in cerebellar FN has never been established. In this study, we investigated the effect of histamine on neuronal firing of cerebellar FN by using slice preparations. Sixty-five FN cells were recorded from 47 cerebellar slices, and a vast majority of the cells responded to histamine stimulation with an excitatory response （58/65, 89.2%）. Perfusing slices with low-Ca^2＋/high-Mg^2＋ medium did not block the histamine-induced excitation （n=10）, supporting a direct postsynaptic action of histamine on the cells. Furthermore, the excitatory effect of histamine on FN neurons was not blocked by selective histamine H1 receptor antagonist triprolidine （n=15） or chlorpheniramine （n=10）, but was effectively suppressed by ranitidine （n=15）, a highly selective histamine H2 receptor antagonist. On the other hand, highly selective histamine H2 receptor agonist dimaprit （n=20） instead of histamine HI receptor agonist 2-pyridylethylamine （n=16） mimicked the excitatory effect of histamine on FN neurons. The dimaprit-induced FN neuronal excitation was effectively antagonized by selective histamine H2 receptor antagonist ranitidine （n=13） but not influenced by selective histamine H1 receptor antagonist triprolidine （n=15）. These results demonstrate that histamine excites cerebellar FN cells via the histamine H2 receptor mechanism and suggest that the hypothalamocerebellar histaminergic fibers may modulate cerebellar FN-mediated sensorimotor integration through their excitatory innervations on FN neurons.