Gene profiling of laser-microdissected brain regions and sub-regions

Sanna, Pietro Paolo; King, Alvin R.; Stap, Lena D. van der; Repunte‐Canonigo, Vez

doi:10.1016/j.brainresprot.2005.04.002

Cited by 17 publications

(8 citation statements)

References 12 publications

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“…Laser microdissection was carried out using a Leica AS LMD as previously described (Sanna et al, 2005). Briefly, anesthetized animals were perfused with an RNA preservative (RNAlater, Ambion) diluted 1:10 in water, brains were rapidly removed and snap frozen in 2-methylbutane and stored at −80°C until laser microdissection (Sanna et al, 2005).…”

Section: Methodsmentioning

confidence: 99%

“…Briefly, anesthetized animals were perfused with an RNA preservative (RNAlater, Ambion) diluted 1:10 in water, brains were rapidly removed and snap frozen in 2-methylbutane and stored at −80°C until laser microdissection (Sanna et al, 2005). Total RNA was prepared from microdissected tissue samples with the Picopure RNA Isolation Kit (Arcturus).…”

Section: Methodsmentioning

confidence: 99%

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Genome-wide gene expression analysis identifies K-ras as a regulator of alcohol intake

et al. 2010

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Adaptations in the anterior cingulate cortex (ACC) have been implicated in alcohol and drug addiction. To identify genes that may contribute to excessive drinking, here we performed microarray analyses in laser microdissected rat ACC after a single or repeated administration of an intoxicating dose of alcohol (3g/kg). Expression of the small G protein K-ras was reduced following both single and repeated alcohol administration. We also observed that voluntary alcohol intake in K-ras heterozygous null mice (K-ras +/− ) did not increased after withdrawal from repeated cycles of intermittent ethanol vapor exposure, unlike in their wild-type littermates. To identify K-ras regulated pathways, we then profiled gene expression in the ACC of K-ras +/− , heterozygous null mice for the K-ras negative regulator Nf1 (Nf1 +/− ) and wild-type mice following repeated administration of an intoxicating dose of alcohol. Pathway analysis showed that alcohol differentially affected various pathways in a K-ras dependent manner -some of which previously shown to be regulated by alcohol -including the insulin/PI3K pathway, the NF-kB, the phosphodiesterases (PDEs) pathway, the Jak/Stat and the adipokine signaling pathways. Altogether, the data implicate K-ras-regulated pathways in the regulation of excessive alcohol drinking after a history of dependence.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Genome-wide gene expression analysis identifies K-ras as a regulator of alcohol intake

et al. 2010

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“…Four adult Sprague Dawley male rats (300 -350 g body weight) were anesthetized with chloral hydrate (35 mg/100 g) and perfused transcardially with 10% RNAlater (Ambion) in 0.1 M PB, pH 7.4, as described previously (Sanna et al, 2005). Brains were immediately removed and frozen in isopentane at Ϫ40°C.…”

Section: Phenotypic Characterization Of Retrograde Labeled Cells By Cmentioning

confidence: 99%

Mesocorticolimbic Glutamatergic Pathway

Yamaguchi

Wang²,

Li³

et al. 2011

Journal of Neuroscience

296

328

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The mesocorticolimbic dopamine (DA) system plays important roles in reward, motivation, learning, memory, and movement. This system arises from the A10 region, comprising the ventral tegmental area and three adjacent midline nuclei (caudal linear nucleus, interfascicular nucleus, and rostral linear nucleus of the raphe). DAergic and GABAergic neurons are intermingled in this region with recently discovered glutamatergic neurons expressing the vesicular glutamate transporter 2 (VGluT2). Here, we show by in situ hybridization and immunohistochemistry that there are two subpopulations of neurons expressing VGluT2 mRNA in the A10 region: (1) a major subpopulation that expresses VGluT2 but lacks tyrosine hydroxylase (TH; VGluT2-only neurons), present in each nucleus of the A10 region, and (2) a smaller subpopulation that coexpresses VGluT2 and TH (VGluT2-TH neurons). By quantitative real-time PCR, we determined the mRNA copy numbers encoding VGluT2 or TH in samples of individual microdissected TH immunoreactive (IR) neurons. Data from both in situ hybridization and from mRNA quantification showed that VGluT2 mRNA is not present in every TH-IR neuron, but restricted to a subset of TH-IR neurons located in the medial portion of the A10 region. By integration of tract tracing, in situ hybridization, and immunohistochemistry, we found that VGluT2-only neurons and VGluT2-TH neurons each innervate both the prefrontal cortex and the nucleus accumbens. These findings establish that in addition to the well-recognized mesocorticolimbic DA-only and GABA-only pathways, there exist parallel mesocorticolimbic glutamate-only and glutamate-DA pathways.

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“…Individual rats received a single sc injection of either the vehicle (mix of 1 vol dimethylsulfoxide and 9 vol sunflower oil) or E2 (50 g/kg body weight) with 10-min shifts. Twenty-four hours later, animals were deeply anesthetized with pentobarbital (40 mg/kg body weight, ip) and perfused transcardially with ice-cold PBS containing 10% RNAlater stabilization reagent (QIAGEN, Hilden, Germany) to preserve RNA integrity (28). Brains were removed, and a 3-mm-thick coronal slice was dissected from the forebrain with two blades positioned at bregma 5.2 and 2.2 in a rat brain matrix.…”

Section: Animal Treatments and Preparation Of The Frontal Cortex For mentioning

confidence: 99%

Gene Expression Profiling Identifies Key Estradiol Targets in the Frontal Cortex of the Rat

et al. 2010

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Estradiol modulates a wide range of neural functions in the frontal cerebral cortex where subsets of neurons express estrogen receptor-alpha and -beta. Through these receptors, estradiol contributes to the maintenance of normal operation of the frontal cortex. During the decline of gonadal hormones, the frequency of neurological and psychiatric disorders increases. To shed light on the etiology of disorders related to declining levels of estrogens, we studied the genomic responses to estradiol. Ovariectomized rats were treated with a sc injection of estradiol. Twenty-four hours later, samples from the frontal cortices were dissected, and their mRNA content was analyzed. One hundred thirty-six estradiol-regulated transcripts were identified on Rat 230 2.0 Expression Array. Of the 136 estrogen-regulated genes, 26 and 36 genes encoded proteins involved in the regulation of transcription and signal transduction, respectively. Thirteen genes were related to the calcium signaling pathway. They comprised five genes coding for neurotransmitter receptors. Transcription of three neuropeptides, including cocaine- and amphetamine-regulated transcript, were up-regulated. Fifty-two genes were selected for validation, and 12 transcriptional changes were confirmed. These results provided evidence that estradiol evokes broad transcriptional response in the cortex. Modulation of key components of the calcium signaling pathway, dopaminergic, serotonergic, and glutamatergic neurotransmission, may explain the influence of estrogens on cognitive function and behavior. Up-regulation of cocaine- and amphetamine-regulated transcript contributes to the neuroprotective effects of estradiol. Identification of estradiol-regulated genes in the frontal cortex helps to understand the pathomechanism of neurological and psychiatric disorders associated with altered levels of estrogens.

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Gene profiling of laser-microdissected brain regions and sub-regions

Cited by 17 publications

References 12 publications

Genome-wide gene expression analysis identifies K-ras as a regulator of alcohol intake

Genome-wide gene expression analysis identifies K-ras as a regulator of alcohol intake

Mesocorticolimbic Glutamatergic Pathway

Gene Expression Profiling Identifies Key Estradiol Targets in the Frontal Cortex of the Rat

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