Comparison of [3H] phosphatidylinositol and [3H] phosphatidylinositol 4,5-bisphosphate hydrolysis in postmortem human brain membranes and characterization of stimulation by dopamine D1 receptors

Mary A. Pacheco, Richard S Jope

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45 Citations (Scopus)

Abstract

Assessing the function of the phosphoinositide signal transduction system in membranes prepared from postmortem human brain by measuring the hydrolysis of exogenous labeled phosphoinositides has been applied to studies of a variety of CNS disorders in recent years. Two issues concerning such studies were addressed in the current investigation: how do [3H]phosphatidylinositol and [3H]phosphatidylinositol 4,5-bisphosphate compare as substrates, and how do dopamine D1 receptors influence phosphoinositide signaling? Comparisons of [3H] phosphatidylinositol and [3H] phosphatidylinositol 4,5-bisphosphate hydrolysis stimulated by guanosine-5'-O-(3-thiotriphosphate)-activated G proteins and by several receptor agonists demonstrated that in most oases each substrate gave similar relative results in membranes prepared from prefrontal cortices of six individuals. However, using optimal assay conditions, [3H]phosphatidylinositol produced a greater signal-to-noise ratio compared with [3H]phosphatidylinositol 4,5-bisphosphate. Dopamine D1 receptors were demonstrated to be directly coupled to phosphoinositide hydrolysis in human brain membranes, and this response was shown to be mediated by the G(q/11) G protein subtype and by the β-subtype of phospholipase C. Therefore, these results demonstrate that [3H]phosphatidylinositol is a suitable substrate to measure phosphoinositide hydrolysis in human brain membranes and that dopamine D1 receptors directly stimulate this signaling system.

Original languageEnglish
Pages (from-to)639-644
Number of pages6
JournalJournal of Neurochemistry
Volume69
Issue number2
StatePublished - Aug 1 1997
Externally publishedYes

Fingerprint

Dopamine D1 Receptors
Phosphatidylinositols
Hydrolysis
Brain
Membranes
GTP-Binding Proteins
Substrates
Guanosine 5'-O-(3-Thiotriphosphate)
Signal transduction
Signal-To-Noise Ratio
Type C Phospholipases
Prefrontal Cortex
Assays
Signal Transduction
Signal to noise ratio

Keywords

  • Dopamine D1 receptor
  • G protein
  • Phosphatidylinositol 4,5-bisphosphate
  • Phosphoinositide
  • Signal transduction

ASJC Scopus subject areas

  • Biochemistry
  • Cellular and Molecular Neuroscience

Cite this

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title = "Comparison of [3H] phosphatidylinositol and [3H] phosphatidylinositol 4,5-bisphosphate hydrolysis in postmortem human brain membranes and characterization of stimulation by dopamine D1 receptors",
abstract = "Assessing the function of the phosphoinositide signal transduction system in membranes prepared from postmortem human brain by measuring the hydrolysis of exogenous labeled phosphoinositides has been applied to studies of a variety of CNS disorders in recent years. Two issues concerning such studies were addressed in the current investigation: how do [3H]phosphatidylinositol and [3H]phosphatidylinositol 4,5-bisphosphate compare as substrates, and how do dopamine D1 receptors influence phosphoinositide signaling? Comparisons of [3H] phosphatidylinositol and [3H] phosphatidylinositol 4,5-bisphosphate hydrolysis stimulated by guanosine-5'-O-(3-thiotriphosphate)-activated G proteins and by several receptor agonists demonstrated that in most oases each substrate gave similar relative results in membranes prepared from prefrontal cortices of six individuals. However, using optimal assay conditions, [3H]phosphatidylinositol produced a greater signal-to-noise ratio compared with [3H]phosphatidylinositol 4,5-bisphosphate. Dopamine D1 receptors were demonstrated to be directly coupled to phosphoinositide hydrolysis in human brain membranes, and this response was shown to be mediated by the G(q/11) G protein subtype and by the β-subtype of phospholipase C. Therefore, these results demonstrate that [3H]phosphatidylinositol is a suitable substrate to measure phosphoinositide hydrolysis in human brain membranes and that dopamine D1 receptors directly stimulate this signaling system.",
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