Performance of mouse neural stem cells as a screening reagent: characterization of PAC1 activity in medium-throughput functional assays

TY - JOUR

T1 - Performance of mouse neural stem cells as a screening reagent

T2 - characterization of PAC1 activity in medium-throughput functional assays

AU - Lodge, Anthony P

AU - Langmead, Christopher J

AU - Daniel, Guillaume

AU - Anderson, Greg W

AU - Werry, Tim D

PY - 2010/2

Y1 - 2010/2

N2 - The self-renewal and phenotypic properties of neural stem cells make them an abundant and more physiologically relevant alternative to recombinant cell lines for drug screens to identify ligands acting at neural targets. Here, the authors use high-throughput phenotypic and signaling assays to test the ability of neural stem cells isolated from postnatal mouse hippocampus (mNSCs) to deliver high-content and physiologically relevant data on native peptide receptor activity. The authors find that mNSCs express PAC1 but not the related VPAC1 and VPAC2 receptors. PAC1 promotes both the proliferation of mNSCs and their differentiation into neuronal-like cells. In addition, the authors show that PAC1 stimulates markedly different extracellular signal-regulated kinase signals in mNSCs than in recombinant CHO-PAC1 cells and is able to couple to Ca(2+) elevation only in CHO-PAC1 cells. These data suggest that G-protein coupling in CHO-PAC1 cells is nonphysiological, which may affect the ligand binding properties of the receptor and thus distort the results of a screen by increasing numbers of false positives/negatives. This work reinforces the emerging pharmacological theory that recombinant cell lines are often inappropriate models of natively expressing primary cells, and the authors conclude that mNSCs are a viable and relevant physiological alternative for use in high-throughput drug screens.

AB - The self-renewal and phenotypic properties of neural stem cells make them an abundant and more physiologically relevant alternative to recombinant cell lines for drug screens to identify ligands acting at neural targets. Here, the authors use high-throughput phenotypic and signaling assays to test the ability of neural stem cells isolated from postnatal mouse hippocampus (mNSCs) to deliver high-content and physiologically relevant data on native peptide receptor activity. The authors find that mNSCs express PAC1 but not the related VPAC1 and VPAC2 receptors. PAC1 promotes both the proliferation of mNSCs and their differentiation into neuronal-like cells. In addition, the authors show that PAC1 stimulates markedly different extracellular signal-regulated kinase signals in mNSCs than in recombinant CHO-PAC1 cells and is able to couple to Ca(2+) elevation only in CHO-PAC1 cells. These data suggest that G-protein coupling in CHO-PAC1 cells is nonphysiological, which may affect the ligand binding properties of the receptor and thus distort the results of a screen by increasing numbers of false positives/negatives. This work reinforces the emerging pharmacological theory that recombinant cell lines are often inappropriate models of natively expressing primary cells, and the authors conclude that mNSCs are a viable and relevant physiological alternative for use in high-throughput drug screens.

KW - Adenosine Triphosphate

KW - Animals

KW - CHO Cells

KW - Calcium

KW - Calcium Signaling

KW - Cell Proliferation

KW - Cricetinae

KW - Cricetulus

KW - Dose-Response Relationship, Drug

KW - Enzyme Activation

KW - Extracellular Signal-Regulated MAP Kinases

KW - Hippocampus

KW - Humans

KW - Mice

KW - Neurons

KW - Phosphorylation

KW - Pituitary Adenylate Cyclase-Activating Polypeptide

KW - Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I

KW - Stem Cells

KW - Time Factors

KW - Vasoactive Intestinal Peptide

U2 - 10.1177/1087057109355468

DO - 10.1177/1087057109355468

M3 - Article

C2 - 20042531

SN - 1087-0571

VL - 15

SP - 159

EP - 168

JO - JOURNAL OF BIOMOLECULAR SCREENING

JF - JOURNAL OF BIOMOLECULAR SCREENING

IS - 2

M1 - N/A

ER -