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The neuroactive steroid pregnenolone sulfate stimulates trafficking of functional N-methyl D-aspartate receptors to the cell surface via a noncanonical, G protein, and Ca2+-dependent mechanism.

ABSTRACT: N-methyl D-aspartate (NMDA) receptors (NMDARs) mediate fast excitatory synaptic transmission and play a critical role in synaptic plasticity associated with learning and memory. NMDAR hypoactivity has been implicated in the pathophysiology of schizophrenia, and clinical studies have revealed reduced negative symptoms of schizophrenia with a dose of pregnenolone that elevates serum levels of the neuroactive steroid pregnenolone sulfate (PregS). This report describes a novel process of delayed-onset potentiation whereby PregS approximately doubles the cell's response to NMDA via a mechanism that is pharmacologically and kinetically distinct from rapid positive allosteric modulation by PregS. The number of functional cell-surface NMDARs in cortical neurons increases 60-100% within 10 minutes of exposure to PregS, as shown by surface biotinylation and affinity purification. Delayed-onset potentiation is reversible and selective for expressed receptors containing the NMDAR subunit subtype 2A (NR2A) or NR2B, but not the NR2C or NR2D, subunits. Moreover, substitution of NR2B J/K helices and M4 domain with the corresponding region of NR2D ablates rapid allosteric potentiation of the NMDA response by PregS but not delayed-onset potentiation. This demonstrates that the initial phase of rapid positive allosteric modulation is not a first step in NMDAR upregulation. Delayed-onset potentiation by PregS occurs via a noncanonical, pertussis toxin-sensitive, G protein-coupled, and Ca(2+)-dependent mechanism that is independent of NMDAR ion channel activation. Further investigation into the sequelae for PregS-stimulated trafficking of NMDARs to the neuronal cell surface may uncover a new target for the pharmacological treatment of disorders in which NMDAR hypofunction has been implicated.

SUBMITTER: Kostakis E

PROVIDER: S-EPMC3716320 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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The neuroactive steroid pregnenolone sulfate stimulates trafficking of functional N-methyl D-aspartate receptors to the cell surface via a noncanonical, G protein, and Ca2+-dependent mechanism.

Kostakis Emmanuel E Smith Conor C Jang Ming-Kuei MK Martin Stella C SC Richards Kyle G KG Russek Shelley J SJ Gibbs Terrell T TT Farb David H DH

Molecular pharmacology 20130528 2

N-methyl D-aspartate (NMDA) receptors (NMDARs) mediate fast excitatory synaptic transmission and play a critical role in synaptic plasticity associated with learning and memory. NMDAR hypoactivity has been implicated in the pathophysiology of schizophrenia, and clinical studies have revealed reduced negative symptoms of schizophrenia with a dose of pregnenolone that elevates serum levels of the neuroactive steroid pregnenolone sulfate (PregS). This report describes a novel process of delayed-ons ...[more]

PMID: 23716622

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Project description:BACKGROUND:Neurosteroids have various physiological and neuropsychopharmacological effects. In addition to the genomic effects of steroids, some neurosteroids modulate several neurotransmitter receptors and channels, such as N-methyl-D-aspartate receptors, gamma-aminobutyric acid type A (GABA(A)) receptors, and sigma(1) receptors, and voltage-gated Ca(2+) and K(+) channels. However, the molecular mechanisms underlying the various effects of neurosteroids have not yet been sufficiently clarified. In the nervous system, inwardly rectifying K(+) (Kir) channels also play important roles in the control of resting membrane potential, cellular excitability and K(+) homeostasis. Among constitutively active Kir2 channels in a major Kir subfamily, Kir2.3 channels are expressed predominantly in the forebrain, a brain area related to cognition, memory, emotion, and neuropsychiatric disorders. METHODOLOGY/PRINCIPAL FINDINGS:The present study examined the effects of various neurosteroids on Kir2.3 channels using the Xenopus oocyte expression assay. In oocytes injected with Kir2.3 mRNA, only pregnenolone sulfate (PREGS), among nine neurosteroids tested, reversibly potentiated Kir2.3 currents. The potentiation effect was concentration-dependent in the micromolar range, and the current-voltage relationship showed inward rectification. However, the potentiation effect of PREGS was not observed when PREGS was applied intracellularly and was not affected by extracellular pH conditions. Furthermore, although Kir1.1, Kir2.1, Kir2.2, and Kir3 channels were insensitive to PREGS, in oocytes injected with Kir2.1/Kir2.3 or Kir2.2/Kir2.3 mRNA, but not Kir2.1/Kir2.2 mRNA, PREGS potentiated Kir currents. These potentiation properties in the concentration-response relationships were less potent than for Kir2.3 channels, suggesting action of PREGS on Kir2.3-containing Kir2 heteromeric channels. CONCLUSIONS/SIGNIFICANCE:The present results suggest that PREGS acts as a positive modulator of Kir2.3 channels. Kir2.3 channel potentiation may provide novel insights into the various effects of PREGS.

Project description:Positive modulation of GABA(A) and antagonism of N-methyl-D-aspartate receptors mediate the discriminative stimulus effects of ethanol. Endogenous neuroactive steroids produce effects similar to ethanol, suggesting that these steroids may modulate ethanol addiction. The four isomers of the functional esters at C-3 of the 3-hydroxy metabolites of 4-pregnene-3,20-dione (progesterone) [allopregnanolone (3alpha,5alpha-P), pregnanolone (3alpha,5beta-P), epiallopregnanolone (3beta,5alpha-P), and epipregnanolone (3beta,5beta-P)], a synthetic analog of steroids modified by endogenous sulfation [pregnanolone hemisuccinate (3alpha,5beta-P HS)], and a structurally similar, adrenally derived steroid [3alpha-hydroxy-5-androstan-17-one (3alpha,5alpha-A, androsterone)] were assessed for ethanol-like discriminative stimulus effects at 30 or 60 min after administration in male (n = 9) and female (n = 8) cynomolgus monkeys (Macaca fascicularis) trained to discriminate 1.0 or 2.0 g/kg ethanol (i.g.) with a 30-min pretreatment interval. The 3alpha-hydroxysteroids completely substituted for ethanol (80% of cases), whereas the 3beta-hydroxysteroids and 3alpha,5beta-P HS rarely substituted for ethanol (6% of cases). There were no sex differences. Compared with monkeys trained to discriminate 2.0 g/kg ethanol, 3alpha,5beta-P and 3alpha,5alpha-A substituted more potently in monkeys trained to discriminate 1.0 g/kg ethanol. Compared with the 5beta-reduced isomer (3alpha,5beta-P), the 5alpha isomer of pregnanolone (3alpha,5alpha-P) substituted for ethanol with 3 to 40-fold greater potency but was least efficacious in female monkeys trained to discriminate 2.0 g/kg ethanol. The data suggest that the discriminative stimulus effects of lower doses (1.0 g/kg) of ethanol are mediated to a greater extent by 3alpha,5beta-P- and 3alpha,5alpha-A-sensitive receptors compared with higher doses (2.0 g/kg). Furthermore, the discriminative stimulus effects of ethanol appear to be mediated by activity at binding sites that are particularly sensitive to 3alpha,5alpha-P.

Dataset Information

The neuroactive steroid pregnenolone sulfate stimulates trafficking of functional N-methyl D-aspartate receptors to the cell surface via a noncanonical, G protein, and Ca2+-dependent mechanism.

Publications

The neuroactive steroid pregnenolone sulfate stimulates trafficking of functional N-methyl D-aspartate receptors to the cell surface via a noncanonical, G protein, and Ca2+-dependent mechanism.

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