Limits...
Attention deficit-hyperactivity disorder suffers from mitochondrial dysfunction

View Article: PubMed Central - PubMed

ABSTRACT

Background: Pathophysiology of attention-deficit hyperactivity disorder (ADHD) is not known, and therefore the present study investigated mitochondrial defects, if any in cybrids created from patients and control population.

Methods: To investigate mitochondrial pathology in ADHD, cybrids cell lines were created from ADHD probands and controls by fusing their platelets with ρ0-cells prepared from SH-SY5Y neuroblastoma cell line. Cellular respiration, oxidative stress, mitochondrial membrane potential and morphology were evaluated employing oxygraph, mitochondria-specific fluorescence staining and evaluation by FACS, and immunocytochemistry. HPLC-electrochemical detection, quantitative RT-PCR and Blue Native PAGE were employed respectively for assays of serotonin, mitochondrial ATPase 6/8 subunits levels and complex V activity.

Results: Significantly low cellular and mitochondrial respiration, ATPase6/8 transcripts levels, mitochondrial complex V activity and loss of mitochondrial membrane potential and elevated oxidative stress were observed in ADHD cybrids. Expression of monoamine oxidizing mitochondrial enzymes, MAO-A and MAO-B levels remained unaffected. Two-fold increase in serotonin level was noted in differentiated cybrid-neurons.

Conclusions: Since cybrids are shown to replicate mitochondrial defects seen in post-mortem brains, these observed defects in ADHD cybrids strongly suggest mitochondrial pathology in this disorder.

General significance: Mitochondrial defects are detected in ADHD cybrids created from patients' platelets, implying bioenergetics crisis in the mitochondria could be a contributory factor for ADHD pathology and/or phenotypes.

No MeSH data available.


Related in: MedlinePlus

Whole cell and mitochondrial state3 respiration, APTase6/8 subunits expression, complex V activity and serotonin levels in control and ADHD cybrids: Whole cell respiration (A), cellular rate of oxygen consumption per 3 × 106 cells (B), mitochondrial State3 respiration (C), and mitochondrial oxygen consumption rate per 3 × 106 cells in ADHD and control cybrids (D). The cDNA prepared from total RNA of ADHD and control cybrids were subjected to qRT-PCR analysis using specific primers to accurately measure the transcript level of ATPase6/8 (E, F). Levels of serotonin in differentiated control and ADHD cybrid-neurons (G). Complex V activity using BN-PAGE, Activity gel (H), Coomassie staining gel (I). Bar diagram shows the difference in complex V activity analysed by Image software (J). (Control samples- 1 to 4 lanes and ADHD samples- 5 to 7 lanes in both the gels, M lane is marker). Data are presented as Mean ± SEM. Values of p* ≤ 0.05 is considered significant.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5121149&req=5

f0015: Whole cell and mitochondrial state3 respiration, APTase6/8 subunits expression, complex V activity and serotonin levels in control and ADHD cybrids: Whole cell respiration (A), cellular rate of oxygen consumption per 3 × 106 cells (B), mitochondrial State3 respiration (C), and mitochondrial oxygen consumption rate per 3 × 106 cells in ADHD and control cybrids (D). The cDNA prepared from total RNA of ADHD and control cybrids were subjected to qRT-PCR analysis using specific primers to accurately measure the transcript level of ATPase6/8 (E, F). Levels of serotonin in differentiated control and ADHD cybrid-neurons (G). Complex V activity using BN-PAGE, Activity gel (H), Coomassie staining gel (I). Bar diagram shows the difference in complex V activity analysed by Image software (J). (Control samples- 1 to 4 lanes and ADHD samples- 5 to 7 lanes in both the gels, M lane is marker). Data are presented as Mean ± SEM. Values of p* ≤ 0.05 is considered significant.

Mentions: Oxygen consumption was monitored in controls and ADHD non-permeabilized (Fig. 3A and B) and permeabilized cells (Fig. 3C and D) employing a sensitive Oxygraph in order to understand the respiratory capability of the cybrids. Significantly reduced rate of respiration found in ADHD cybrids, as well as in the mitochondria of these disease-cybrids, pointed to a significant loss of mitochondrial functions in both these diseases. Lower respiration rate could be resulting from the considerable loss in the ATPase 6/8 transcript levels in ADHD cybrids revealed in quantitative PCR analysis as seen in the present study (Fig. 3E and F).


Attention deficit-hyperactivity disorder suffers from mitochondrial dysfunction
Whole cell and mitochondrial state3 respiration, APTase6/8 subunits expression, complex V activity and serotonin levels in control and ADHD cybrids: Whole cell respiration (A), cellular rate of oxygen consumption per 3 × 106 cells (B), mitochondrial State3 respiration (C), and mitochondrial oxygen consumption rate per 3 × 106 cells in ADHD and control cybrids (D). The cDNA prepared from total RNA of ADHD and control cybrids were subjected to qRT-PCR analysis using specific primers to accurately measure the transcript level of ATPase6/8 (E, F). Levels of serotonin in differentiated control and ADHD cybrid-neurons (G). Complex V activity using BN-PAGE, Activity gel (H), Coomassie staining gel (I). Bar diagram shows the difference in complex V activity analysed by Image software (J). (Control samples- 1 to 4 lanes and ADHD samples- 5 to 7 lanes in both the gels, M lane is marker). Data are presented as Mean ± SEM. Values of p* ≤ 0.05 is considered significant.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5121149&req=5

f0015: Whole cell and mitochondrial state3 respiration, APTase6/8 subunits expression, complex V activity and serotonin levels in control and ADHD cybrids: Whole cell respiration (A), cellular rate of oxygen consumption per 3 × 106 cells (B), mitochondrial State3 respiration (C), and mitochondrial oxygen consumption rate per 3 × 106 cells in ADHD and control cybrids (D). The cDNA prepared from total RNA of ADHD and control cybrids were subjected to qRT-PCR analysis using specific primers to accurately measure the transcript level of ATPase6/8 (E, F). Levels of serotonin in differentiated control and ADHD cybrid-neurons (G). Complex V activity using BN-PAGE, Activity gel (H), Coomassie staining gel (I). Bar diagram shows the difference in complex V activity analysed by Image software (J). (Control samples- 1 to 4 lanes and ADHD samples- 5 to 7 lanes in both the gels, M lane is marker). Data are presented as Mean ± SEM. Values of p* ≤ 0.05 is considered significant.
Mentions: Oxygen consumption was monitored in controls and ADHD non-permeabilized (Fig. 3A and B) and permeabilized cells (Fig. 3C and D) employing a sensitive Oxygraph in order to understand the respiratory capability of the cybrids. Significantly reduced rate of respiration found in ADHD cybrids, as well as in the mitochondria of these disease-cybrids, pointed to a significant loss of mitochondrial functions in both these diseases. Lower respiration rate could be resulting from the considerable loss in the ATPase 6/8 transcript levels in ADHD cybrids revealed in quantitative PCR analysis as seen in the present study (Fig. 3E and F).

View Article: PubMed Central - PubMed

ABSTRACT

Background: Pathophysiology of attention-deficit hyperactivity disorder (ADHD) is not known, and therefore the present study investigated mitochondrial defects, if any in cybrids created from patients and control population.

Methods: To investigate mitochondrial pathology in ADHD, cybrids cell lines were created from ADHD probands and controls by fusing their platelets with ρ0-cells prepared from SH-SY5Y neuroblastoma cell line. Cellular respiration, oxidative stress, mitochondrial membrane potential and morphology were evaluated employing oxygraph, mitochondria-specific fluorescence staining and evaluation by FACS, and immunocytochemistry. HPLC-electrochemical detection, quantitative RT-PCR and Blue Native PAGE were employed respectively for assays of serotonin, mitochondrial ATPase 6/8 subunits levels and complex V activity.

Results: Significantly low cellular and mitochondrial respiration, ATPase6/8 transcripts levels, mitochondrial complex V activity and loss of mitochondrial membrane potential and elevated oxidative stress were observed in ADHD cybrids. Expression of monoamine oxidizing mitochondrial enzymes, MAO-A and MAO-B levels remained unaffected. Two-fold increase in serotonin level was noted in differentiated cybrid-neurons.

Conclusions: Since cybrids are shown to replicate mitochondrial defects seen in post-mortem brains, these observed defects in ADHD cybrids strongly suggest mitochondrial pathology in this disorder.

General significance: Mitochondrial defects are detected in ADHD cybrids created from patients' platelets, implying bioenergetics crisis in the mitochondria could be a contributory factor for ADHD pathology and/or phenotypes.

No MeSH data available.


Related in: MedlinePlus