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Constitutive overexpression of muscarinic receptors leads to vagal hyperreactivity.

Livolsi A, Niederhoffer N, Dali-Youcef N, Mokni W, Olexa-Zorn C, Gies JP, Marcellin L, Feldman J, Bousquet P - PLoS ONE (2010)

Bottom Line: Sequencing of the coding fragment of the M(2) receptor gene revealed a single nucleotide mutation in 83% of hyperreactive animals, possibly contributing for the transcript overexpression.This phenomenon might represent a compensatory consequence of muscarinic receptors overexpression.The results suggest that cardiac muscarinic receptor overexpression plays a critical role in the development of vagal hyperreactivity, whereas AchE hyperactivity appears as a compensatory consequence of it.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Neurobiologie et Pharmacologie Cardiovasculaire (EA 4438), Université de Strasbourg, Strasbourg, France.

ABSTRACT

Background: Alterations in muscarinic receptor expression and acetylcholinesterase (AchE) activity have been observed in tissues from Sudden Infant Death Syndrome (SIDS). Vagal overactivity has been proposed as a possible cause of SIDS as well as of vasovagal syncopes. The aim of the present study was to seek whether muscarinic receptor overexpression may be the underlying mechanism of vagal hyperreactivity. Rabbits with marked vagal pauses following injection of phenylephrine were selected and crossed to obtain a vagal hyperreactive strain. The density of cardiac muscarinic receptors and acetylcholinesterase (AchE) gene expression were assessed. Blood markers of the observed cardiac abnormalities were also sought.

Methodology/principal findings: Cardiac muscarinic M(2) and M(3) receptors were overexpressed in hyperreactive rabbits compared to control animals (2.3-fold and 2.5-fold, respectively) and the severity of the phenylephrine-induced bradycardia was correlated with their densities. A similar overexpression of M(2) receptors was observed in peripheral mononuclear white blood cells, suggesting that cardiac M(2) receptor expression can be inferred with high confidence from measurements in blood cells. Sequencing of the coding fragment of the M(2) receptor gene revealed a single nucleotide mutation in 83% of hyperreactive animals, possibly contributing for the transcript overexpression. Significant increases in AchE expression and activity were also assessed (AchE mRNA amplification ratio of 3.6 versus normal rabbits). This phenomenon might represent a compensatory consequence of muscarinic receptors overexpression. Alterations in M(2) receptor and AchE expression occurred between the 5th and the 7th week of age, a critical period also characterized by a higher mortality rate of hyperreactive rabbits (52% in H rabbits versus 13% in normal rabbits) and preceeded the appearance of functional disorders.

Conclusions/significance: The results suggest that cardiac muscarinic receptor overexpression plays a critical role in the development of vagal hyperreactivity, whereas AchE hyperactivity appears as a compensatory consequence of it. Since similar vagal disorders were observed recently by us in SIDS, muscarinic receptor overexpression could become a marker of risk of vasovagal syncopes and SIDS.

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M2 muscarinic receptor gene expression in peripheral mononuclear white blood cells from normal (N) and vagal hyperreactive (H) rabbits.R-R intervals were measured in conscious rabbits challenged with PNE 500 µg kg−1 following the procedure described in Material and Methods. M2 muscarinic gene expression was assessed in peripheral mononuclear white blood cells by quantitative RT-PCR and normalized to the rabbit 18S housekeeping gene. Values in (a) show amplification ratio calculated according to the 2−ΔΔCt method of 9 (N) and 16 (H) experiments. In (b), each symbol represents one animal; ΔCt M2-18S corresponds to the number of amplification cycles needed to detect M2 fluorescence standardized to 18S; thus, the lower the ΔCt M2-18S, the greater M2 mRNA initial quantity. *: P<0.0001 versus N.
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pone-0015618-g002: M2 muscarinic receptor gene expression in peripheral mononuclear white blood cells from normal (N) and vagal hyperreactive (H) rabbits.R-R intervals were measured in conscious rabbits challenged with PNE 500 µg kg−1 following the procedure described in Material and Methods. M2 muscarinic gene expression was assessed in peripheral mononuclear white blood cells by quantitative RT-PCR and normalized to the rabbit 18S housekeeping gene. Values in (a) show amplification ratio calculated according to the 2−ΔΔCt method of 9 (N) and 16 (H) experiments. In (b), each symbol represents one animal; ΔCt M2-18S corresponds to the number of amplification cycles needed to detect M2 fluorescence standardized to 18S; thus, the lower the ΔCt M2-18S, the greater M2 mRNA initial quantity. *: P<0.0001 versus N.

Mentions: M2 mRNA expression level was about 10 times higher in vagal hyperreactive rabbits compared to that observed in normal rabbits (Fig. 2a). Compared to normal animals, M2 mRNA relative quantities were increased in all rabbits displaying baroreflex dysfunction and vagal hyperreactivity (Fig. 2b).


Constitutive overexpression of muscarinic receptors leads to vagal hyperreactivity.

Livolsi A, Niederhoffer N, Dali-Youcef N, Mokni W, Olexa-Zorn C, Gies JP, Marcellin L, Feldman J, Bousquet P - PLoS ONE (2010)

M2 muscarinic receptor gene expression in peripheral mononuclear white blood cells from normal (N) and vagal hyperreactive (H) rabbits.R-R intervals were measured in conscious rabbits challenged with PNE 500 µg kg−1 following the procedure described in Material and Methods. M2 muscarinic gene expression was assessed in peripheral mononuclear white blood cells by quantitative RT-PCR and normalized to the rabbit 18S housekeeping gene. Values in (a) show amplification ratio calculated according to the 2−ΔΔCt method of 9 (N) and 16 (H) experiments. In (b), each symbol represents one animal; ΔCt M2-18S corresponds to the number of amplification cycles needed to detect M2 fluorescence standardized to 18S; thus, the lower the ΔCt M2-18S, the greater M2 mRNA initial quantity. *: P<0.0001 versus N.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3008725&req=5

pone-0015618-g002: M2 muscarinic receptor gene expression in peripheral mononuclear white blood cells from normal (N) and vagal hyperreactive (H) rabbits.R-R intervals were measured in conscious rabbits challenged with PNE 500 µg kg−1 following the procedure described in Material and Methods. M2 muscarinic gene expression was assessed in peripheral mononuclear white blood cells by quantitative RT-PCR and normalized to the rabbit 18S housekeeping gene. Values in (a) show amplification ratio calculated according to the 2−ΔΔCt method of 9 (N) and 16 (H) experiments. In (b), each symbol represents one animal; ΔCt M2-18S corresponds to the number of amplification cycles needed to detect M2 fluorescence standardized to 18S; thus, the lower the ΔCt M2-18S, the greater M2 mRNA initial quantity. *: P<0.0001 versus N.
Mentions: M2 mRNA expression level was about 10 times higher in vagal hyperreactive rabbits compared to that observed in normal rabbits (Fig. 2a). Compared to normal animals, M2 mRNA relative quantities were increased in all rabbits displaying baroreflex dysfunction and vagal hyperreactivity (Fig. 2b).

Bottom Line: Sequencing of the coding fragment of the M(2) receptor gene revealed a single nucleotide mutation in 83% of hyperreactive animals, possibly contributing for the transcript overexpression.This phenomenon might represent a compensatory consequence of muscarinic receptors overexpression.The results suggest that cardiac muscarinic receptor overexpression plays a critical role in the development of vagal hyperreactivity, whereas AchE hyperactivity appears as a compensatory consequence of it.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Neurobiologie et Pharmacologie Cardiovasculaire (EA 4438), Université de Strasbourg, Strasbourg, France.

ABSTRACT

Background: Alterations in muscarinic receptor expression and acetylcholinesterase (AchE) activity have been observed in tissues from Sudden Infant Death Syndrome (SIDS). Vagal overactivity has been proposed as a possible cause of SIDS as well as of vasovagal syncopes. The aim of the present study was to seek whether muscarinic receptor overexpression may be the underlying mechanism of vagal hyperreactivity. Rabbits with marked vagal pauses following injection of phenylephrine were selected and crossed to obtain a vagal hyperreactive strain. The density of cardiac muscarinic receptors and acetylcholinesterase (AchE) gene expression were assessed. Blood markers of the observed cardiac abnormalities were also sought.

Methodology/principal findings: Cardiac muscarinic M(2) and M(3) receptors were overexpressed in hyperreactive rabbits compared to control animals (2.3-fold and 2.5-fold, respectively) and the severity of the phenylephrine-induced bradycardia was correlated with their densities. A similar overexpression of M(2) receptors was observed in peripheral mononuclear white blood cells, suggesting that cardiac M(2) receptor expression can be inferred with high confidence from measurements in blood cells. Sequencing of the coding fragment of the M(2) receptor gene revealed a single nucleotide mutation in 83% of hyperreactive animals, possibly contributing for the transcript overexpression. Significant increases in AchE expression and activity were also assessed (AchE mRNA amplification ratio of 3.6 versus normal rabbits). This phenomenon might represent a compensatory consequence of muscarinic receptors overexpression. Alterations in M(2) receptor and AchE expression occurred between the 5th and the 7th week of age, a critical period also characterized by a higher mortality rate of hyperreactive rabbits (52% in H rabbits versus 13% in normal rabbits) and preceeded the appearance of functional disorders.

Conclusions/significance: The results suggest that cardiac muscarinic receptor overexpression plays a critical role in the development of vagal hyperreactivity, whereas AchE hyperactivity appears as a compensatory consequence of it. Since similar vagal disorders were observed recently by us in SIDS, muscarinic receptor overexpression could become a marker of risk of vasovagal syncopes and SIDS.

Show MeSH
Related in: MedlinePlus