laser-capture-microdissection-demonstration-isolation-individual

Laser Capture Microdissection - A Demonstration of the Isolation of Individual Dopamine Neurons and the Entire Ventral Tegmental Area

Laser capture microdissection (LCM) is used to isolate a concentrated population of individual cells or precise anatomical regions of tissue from tissue ...

Mississippi State University College of Veterinary Medicine

Choline, a dietary compound present in many foods, has recently been classified as an essential nutrient for humans. There is evidence from animal models that the availability of choline during the prenatal period influences neural and cognitive development. Here we report that choline supplementation during a 6 d gestational period protects against neurodegeneration in the posterior cingulate and retrosplenial cortices of female adolescent rats produced by peripheral administration of the NMDA receptor antagonist dizocilpine (MK-801). These data show that availability of a single nutrient, choline, during a brief period of prenatal development diminishes vulnerability to neurotoxicity in adolescent offspring.

Prenatal choline supplementation protects against postnatal neurotoxicity.

The availability of choline during the prenatal period influences neural and cognitive development. Here we report that choline supplementation during a six-day gestational period protects against neurodegeneration in the posterior cingulate and retrosplenial cortices of adult female rats produced by systemic administration of the N-methyl-D-aspartate receptor antagonist dizocilpine (MK-801). These data show that availability of choline during a brief prenatal period diminishes vulnerability to neurotoxicity in adult offspring.

Prenatal dietary choline availability alters postnatal neurotoxic vulnerability in the adult rat.

The neurodevelopmental effects of two organophosphorus (OP) insecticides, chlorpyrifos (CPS) and methyl parathion (MPS), on cholinesterase (ChE) activity and muscarinic acetylcholine receptor (mAChR) binding were investigated in neonatal rat brain. Animals were orally gavaged using an incremental dosing regimen from postnatal day 1 (PND1) until PND8 with a low, medium, and high dosage for both CPS and MPS. On PND4 and PND8, ChE activity was measured in whole brain while the total and subtype densities of mAChRs were measured in three brain sections: area anterior to optic chiasma (anterior forebrain), area from the optic chiasma to the medulla/pons (posterior forebrain); and the medulla/pons excluding the cerebellum. The ligands 3H-pirenzepine, 3H-AF-DX 384, 3H-4-DAMP, and 3H-QNB were used to measure the maximal binding of the M1, M2/M4, and M3 subtypes and total mAChR receptors, respectively. In the anterior and the posterior forebrain, the levels of all mAChRs nearly doubled from PND4 to PND8, while in the medulla/pons, M1- and M3-subtype mAChR densities were low and did not increase and M2/M4 subtype and total mAChR slightly increased from PND4 to PND8. Reduction of ChE activity and mAChR binding by CPS or MPS was more evident in rats at PND8 than at PND4. With respect to mAChR binding, the greatest effects were observed in the medulla/pons and the least effects were observed in the posterior region of the forebrain. These results demonstrate that OPs exert adverse effects on rat central nervous system development through the cholinergic system in an age- and region-dependent manner.

Altered muscarinic acetylcholine receptor subtype binding in neonatal rat brain following exposure to chlorpyrifos or methyl parathion.

The NR4A transcription factors NR4A1, NR4A2, and NR4A3 (also known as Nur77, Nurr1, and Nor1, respectively) share similar DNA-binding properties and have been implicated in regulation of dopamine neurotransmission genes. Our current hypothesis is that NR4A gene expression is regulated by dopamine neuron activity and that induction of NR4A genes will increase expression of dopamine neurotransmission genes. Eticlopride and γ-butyrolactone (GBL) were used in wild-type (+/+) and Nurr1-null heterozygous (+/-) mice to determine the mechanism(s) regulating Nur77 and Nurr1 expression. Laser capture microdissection and real-time PCR was used to measure Nurr1 and Nur77 mRNA levels in the ventral tegmental area (VTA). Nur77 expression was significantly elevated 1 h after both GBL (twofold) and eticlopride (fourfold). In contrast, GBL significantly decreased Nurr1 expression in both genotypes, while eticlopride significantly increased Nurr1 expression only in the +/+ mice. In a separate group of mice, haloperidol injection significantly elevated Nur77 and Nor1, but not Nurr1 mRNA in the VTA within 1 h and significantly increased tyrosine hydroxylase (TH) and dopamine transporter (DAT) mRNA expression by 4 h. These data demonstrate that the NR4A genes are dynamically regulated in dopamine neurons with maintenance of Nurr1 expression requiring dopamine neuron activity while both attenuation of dopamine autoreceptors activation and dopamine neuronal activity combining to induce Nur77 expression. Additionally, these data suggest that induction of NR4A genes could regulate TH and DAT expression and ultimately regulate dopamine neurotransmission.

NR4A gene expression is dynamically regulated in the ventral tegmental area dopamine neurons and is related to expression of dopamine neurotransmission genes.

Shirley X. Guo-Ross

Department of Basic Sciences

Mississippi State University College of Veterinary Medicine

Laser Capture Microdissection - A Demonstration of the Isolation of Individual Dopamine Neurons and the Entire Ventral Tegmental Area

Shirley X. Guo-Ross

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Basic Sciences

Mississippi State University College of Veterinary Medicine

Laser Capture Microdissection - A Demonstration of the Isolation of Individual Dopamine Neurons and the Entire Ventral Tegmental Area

Department of Basic Sciences