the major clinical symptoms of migraine include moderate - to - severe pulsatile headaches , often accompanied by nausea and vomiting .
optical and acoustic stimuli , as well as some daily activities , are known to exacerbate migraine headaches , whereas a quiet environment and plenty of rest can relieve the headaches .
valproate is a broad - spectrum antiepileptic drug that is used to over one - third of epileptic patients .
valproate has also been approved by the fda for the treatment of neuralgia and bipolar disorders .
valproate can exert a neuroprotective effect and modulates neuronal differentiation and survival , as well as long - term neuroplasticity .
in addition , valproate can protect the cortical neurons from spontaneous cell death and improve the behavioral outcomes .
valproate is effective in relieving headaches and is proposed to be a potential anti - migraine drug [ 69 ] .
valproate can also be used in the treatment of headaches from medication overuse   and as a prophylaxis of episodic migraine .
mitochondria are important sites of energy metabolism , especially in the high energy - consuming organs .
it has been reported that mitochondrial function is often impaired in patients with migraine , and mitochondrial dysfunction is thus implicated in the pathophysiology of migraine .
valproate can be used to prevent and mitigate migraine , but the underlying mechanism remains unclear . in the present study , we explored the effect of valproate on migraine and the underlying mechanism in a rat model of nitroglycerin - induced trigeminovascular activation .
valproate was found to attenuate nitroglycerin - induced trigeminovascular activation in rats by preserving the mitochondrial function .
male sprague - dawley rats ( 8 weeks old , weighing around 200 g ) were obtained from liaoning changsheng biotechnology co. , ltd .
( benxi , china ) and housed in a controlled environment ( 232c , 505% relative humidity , and 12 h : 12 h light : dark cycles ) .
forty rats were randomly divided into 4 groups ( control , model , model+vp - l , and model+vp - h ) .
rats in the model+vp - l group and model+vp - h group received a low dose of valproate ( 100 mg / kg in saline ) ( dalian meilun biotech co. , ltd . , dalian , china ) or a high dose of valproate ( 200 mg / kg in saline ) every day by intraperitoneal injection for 5 days .
rats in the control and model groups received an equal volume of saline . on the sixth day
, rats in the model , model+vp - l , and model+vp - h groups received an intraperitoneal injection of nitroglycerin ( 10 mg / kg in saline ) ( beijing yimin pharmaceutical co. , ltd . , beijing , china ) , and rats in the control group received an equal volume of saline .
four hours after nitroglycerin injection , the peripheral blood was collected and the levels of 5-hydroxytryptamine ( 5-ht ) and nitric oxide ( no ) were measured using a 5-ht elisa kit ( uscn , wuhan , china ) and a total no assay kit ( beyotime , haimen , china ) respectively .
the rats were then sacrificed , and the spinal trigeminal nucleus was obtained for subsequent experiments .
the care and treatment of the animals was approved by the animal ethics committee of the first affiliated hospital of harbin medical university .
dna was extracted from rat spinal trigeminal nucleus using a dna extraction kit ( tiangen , beijing , china ) according to the manufacturer s instructions .
the mtdna copy number was measured by quantitative real - time pcr according to the methods described previously [ 1417 ] , with the following primers : dn1 forward primer : 5-ttatcctcttatccgtcctc-3 ; dn1 reverse primer : 5-tgttaagtcgaagggagc-3 ; -actin forward primer : 5-agggaaatcgtgcgtgac-3 ; -actin reverse primer : 5-aggaaggaaggctggaag-3. the relative mtdna copy number was calculated using the 2 method .
proteins in the spinal trigeminal nucleus were extracted using a total protein extraction kit ( wanleibio , shenyang , china ) .
after measurement of the protein concentration with a bca protein assay kit ( wanleibio ) , an equal amount of proteins from each group was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis .
subsequently , the separated proteins were transferred onto polyvinylidene fluoride membranes ( millipore , bedford , ma ) , followed by blocking with 5% skim milk . after washing with tbst
, the membranes were incubated overnight at 4c with primary antibodies against peroxisome proliferator - activated receptor- ( pparg ) ( 1:400 , bioss , beijing , china ) , peroxisome proliferator - activated receptor- coactivator-1 ( pgc-1 ) ( 1:1000 , abcam , cambridge , uk ) , mitochondrial transcription factor a ( tfam ) ( 1:2000 , abcam ) , b - cell lymphoma-2 ( bcl-2 ) , bcl-2-associated x protein ( bax ) ( 1:400 , boster , wuhan , china ) , and -actin ( 1:1000 , santa cruz , dallas , tx ) .
thereafter , the membranes were washed with tbst and then incubated with the corresponding horseradish peroxidase - labeled secondary antibody ( 1:5000 , wanleibio ) at 37c for 45 min .
the membranes were washed with tbst again and the signals were detected with an ecl detection system ( wanleibio ) .
the gray intensity of the target proteins was analyzed with gel - pro - analyzer software , and the relative protein level was calculated using -actin as the internal reference .
the spinal trigeminal nucleus was homogenized in 9 volumes of pbs . after freezing and thawing 3 times ,
the protein concentration in the supernatant was measured with a bca protein assay kit , and the atp level was determined with an atp assay kit ( jianchengbio , nanjing , china ) according to the manufacturer s protocol . the protein concentration in the supernatant of spinal trigeminal nucleus homogenate was determined , and the activity of cytochrome c oxidase in the supernatant was detected with a rat cytochrome c oxidase assay kit ( whb , shanghai , china ) according to the manufacturer s instructions .
after measurement of protein concentration , the supernatant of tissue homogenate was used for determining the ros level using an ros assay kit ( jianchengbio ) according to the protocol .
the mitochondrial membrane potential was assessed by staining with the jc-1 probe , followed by fluorescence microscopy or flow cytometry .
the cells were incubated with the jc-1 working solution from the jc-1 kit ( keygen , nanjing , china ) at 37c for 20 min .
the cell images were captured under a fluorescence microscope ( olympus , tokyo , japan ) , and the fluorescence signals were quantitated by flow cytometry ( bd , franklin lakes , nj ) .
the differences between the control group and the model group were analyzed using student s t test , and differences between the model group , model+vp - l group , and model+vp - h group were analyzed using one - way analysis of variance followed by bonferroni s multiple comparison test .
male sprague - dawley rats ( 8 weeks old , weighing around 200 g ) were obtained from liaoning changsheng biotechnology co. , ltd .
( benxi , china ) and housed in a controlled environment ( 232c , 505% relative humidity , and 12 h : 12 h light : dark cycles ) .
forty rats were randomly divided into 4 groups ( control , model , model+vp - l , and model+vp - h ) .
rats in the model+vp - l group and model+vp - h group received a low dose of valproate ( 100 mg / kg in saline ) ( dalian meilun biotech co. , ltd . , dalian , china ) or a high dose of valproate ( 200 mg / kg in saline ) every day by intraperitoneal injection for 5 days .
rats in the control and model groups received an equal volume of saline . on the sixth day
, rats in the model , model+vp - l , and model+vp - h groups received an intraperitoneal injection of nitroglycerin ( 10 mg / kg in saline ) ( beijing yimin pharmaceutical co. , ltd . , beijing , china ) , and rats in the control group received an equal volume of saline .
four hours after nitroglycerin injection , the peripheral blood was collected and the levels of 5-hydroxytryptamine ( 5-ht ) and nitric oxide ( no ) were measured using a 5-ht elisa kit ( uscn , wuhan , china ) and a total no assay kit ( beyotime , haimen , china ) respectively .
the rats were then sacrificed , and the spinal trigeminal nucleus was obtained for subsequent experiments .
the care and treatment of the animals was approved by the animal ethics committee of the first affiliated hospital of harbin medical university .
dna was extracted from rat spinal trigeminal nucleus using a dna extraction kit ( tiangen , beijing , china ) according to the manufacturer s instructions .
the mtdna copy number was measured by quantitative real - time pcr according to the methods described previously [ 1417 ] , with the following primers : dn1 forward primer : 5-ttatcctcttatccgtcctc-3 ; dn1 reverse primer : 5-tgttaagtcgaagggagc-3 ; -actin forward primer : 5-agggaaatcgtgcgtgac-3 ; -actin reverse primer : 5-aggaaggaaggctggaag-3. the relative mtdna copy number was calculated using the 2 method .
proteins in the spinal trigeminal nucleus were extracted using a total protein extraction kit ( wanleibio , shenyang , china ) .
after measurement of the protein concentration with a bca protein assay kit ( wanleibio ) , an equal amount of proteins from each group was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis .
subsequently , the separated proteins were transferred onto polyvinylidene fluoride membranes ( millipore , bedford , ma ) , followed by blocking with 5% skim milk . after washing with tbst
, the membranes were incubated overnight at 4c with primary antibodies against peroxisome proliferator - activated receptor- ( pparg ) ( 1:400 , bioss , beijing , china ) , peroxisome proliferator - activated receptor- coactivator-1 ( pgc-1 ) ( 1:1000 , abcam , cambridge , uk ) , mitochondrial transcription factor a ( tfam ) ( 1:2000 , abcam ) , b - cell lymphoma-2 ( bcl-2 ) , bcl-2-associated x protein ( bax ) ( 1:400 , boster , wuhan , china ) , and -actin ( 1:1000 , santa cruz , dallas , tx ) .
thereafter , the membranes were washed with tbst and then incubated with the corresponding horseradish peroxidase - labeled secondary antibody ( 1:5000 , wanleibio ) at 37c for 45 min .
the membranes were washed with tbst again and the signals were detected with an ecl detection system ( wanleibio ) .
the gray intensity of the target proteins was analyzed with gel - pro - analyzer software , and the relative protein level was calculated using -actin as the internal reference .
the spinal trigeminal nucleus was homogenized in 9 volumes of pbs . after freezing and thawing 3 times ,
the protein concentration in the supernatant was measured with a bca protein assay kit , and the atp level was determined with an atp assay kit ( jianchengbio , nanjing , china ) according to the manufacturer s protocol .
the protein concentration in the supernatant of spinal trigeminal nucleus homogenate was determined , and the activity of cytochrome c oxidase in the supernatant was detected with a rat cytochrome c oxidase assay kit ( whb , shanghai , china ) according to the manufacturer s instructions .
after measurement of protein concentration , the supernatant of tissue homogenate was used for determining the ros level using an ros assay kit ( jianchengbio ) according to the protocol .
the mitochondrial membrane potential was assessed by staining with the jc-1 probe , followed by fluorescence microscopy or flow cytometry .
the cells were incubated with the jc-1 working solution from the jc-1 kit ( keygen , nanjing , china ) at 37c for 20 min .
the cell images were captured under a fluorescence microscope ( olympus , tokyo , japan ) , and the fluorescence signals were quantitated by flow cytometry ( bd , franklin lakes , nj ) .
the differences between the control group and the model group were analyzed using student s t test , and differences between the model group , model+vp - l group , and model+vp - h group were analyzed using one - way analysis of variance followed by bonferroni s multiple comparison test .
after induction of trigeminovascular activation with nitroglycerin , the number of scratching behaviors was recorded . as shown in figure 1a ,
the number of scratching behaviors in the model group was significantly higher than that in the control group . compared with the model group ,
the number of scratching behaviors was decreased in the model group rats with valproate pre - treatment .
consistent with the number of scratching behaviors , the 5-ht level was decreased and the no level was increased in the model group compared with the control group , and the nitroglycerin - induced changes were minimized by valproate ( figure 1b , 1c ) .
these results demonstrate that treatment with valproate attenuates nitroglycerin - induced trigeminovascular activation in rats .
the mtdna copy number was measured in the rat model with or without valproate treatment .
compared with the control group , the mtdna copy number in the model group was decreased ( figure 2a ) . a low dose of valproate significantly attenuated nitroglycerin - induced reduction of mtdna copy number , and a high dose of valproate preserved the mtdna copy number to nearly normal level ( figure 2a ) .
to investigate the effect of valproate on the biogenesis of mitochondria , the protein levels of pgc-1 , tfam , and pparg were detected by western blot analysis .
as shown in figure 2 , the protein levels of pgc-1 , tfam , and pparg were lower in the model group in comparison with the control group ( figure 2b2d ) , but nitroglycerin - induced reduction in the expression of pgc-1 , tfam , and pparg was inhibited by valproate .
the level of atp in the spinal trigeminal nucleus was measured in the nitroglycerin - induced trigeminovascular activation rat models with or without valproate treatment .
the atp level was lowered in the model group , indicating the dysfunction of mitochondrial energy metabolism .
compared with the model group , a low dose of valproate elevated the atp level , which was further increased by a high dose of valproate ( figure 3a ) .
these results suggest that nitroglycerin - induced dysfunction of mitochondrial energy metabolism in the spinal trigeminal nucleus is alleviated by valproate .
the mitochondrial membrane potential is crucial for energy metabolism in the mitochondria . in this study ,
as shown in figure 3b , almost all the jc-1 probes in the control group showed red fluorescence , which indicated a high mitochondrial membrane potential .
in contrast , the jc-1 probes in the model group showed green fluorescence , indicating a low mitochondrial membrane potential in the nitroglycerin - induced trigeminovascular activation rat model .
after treatment with a low dose of valproate , the jc-1 probes showed more red fluorescence and less green fluorescence , while the jc-1 probes in the model+vp - h group showed almost full red fluorescence ( figure 3b ) .
these results indicate that valproate treatment maintains the mitochondrial membrane potential in nitroglycerin - induced trigeminovascular activation .
consistent with the observations by fluorescence microscopy , the mitochondrial membrane potential in the model group was significantly decreased compared with the control group .
however , treatment with valproate prevented nitroglycerin - induced reduction of mitochondrial membrane potential ( figure 3c , 3d ) .
these results demonstrate that valproate preserves the mitochondrial membrane potential in the spinal trigeminal nucleus during nitroglycerin - induced trigeminovascular activation .
bax and bcl-2 have close relationships with the opening of the mitochondrial permeability transition pore . here , the protein levels of bax and bcl-2 were detected by western blot . in the model group ,
the protein level of bax was higher than that in the control group ( figure 4a ) , and the protein level of bcl-2 was lower than that in the control group ( figure 4b ) .
in contrast , treatment with valproate decreased the level of bax and increased the level of bcl-2 in the nitroglycerin - treated rats ( figure 4a , 4b ) .
these results provide molecular evidence for the hypothesis that valproate modulates the mitochondria energy metabolism in subjects with migraine .
the activity of cytochrome c oxidase was also examined in our study . as shown in figure 4c ,
the cytochrome c oxidase activity in the model group was decreased significantly as compared with the control group .
compared with the model group , valproate dose - dependently increased the activity of cytochrome c oxidase in the nitroglycerin - treated rats ( figure 4c ) .
consistent with the disrupted mitochondrial energy metabolism in nitroglycerin - treated rats , a significantly increased ros level was observed in the model group ( figure 4d ) .
treatment with valproate significantly attenuated nitroglycerin - induced ros elevation in the spinal trigeminal nucleus .
after induction of trigeminovascular activation with nitroglycerin , the number of scratching behaviors was recorded . as shown in figure 1a ,
the number of scratching behaviors in the model group was significantly higher than that in the control group . compared with the model group ,
the number of scratching behaviors was decreased in the model group rats with valproate pre - treatment .
consistent with the number of scratching behaviors , the 5-ht level was decreased and the no level was increased in the model group compared with the control group , and the nitroglycerin - induced changes were minimized by valproate ( figure 1b , 1c ) .
these results demonstrate that treatment with valproate attenuates nitroglycerin - induced trigeminovascular activation in rats .
the mtdna copy number was measured in the rat model with or without valproate treatment . compared with the control group ,
the mtdna copy number in the model group was decreased ( figure 2a ) . a low dose of valproate significantly attenuated nitroglycerin - induced reduction of mtdna copy number , and a high dose of valproate preserved the mtdna copy number to nearly normal level ( figure 2a ) .
to investigate the effect of valproate on the biogenesis of mitochondria , the protein levels of pgc-1 , tfam , and pparg were detected by western blot analysis . as shown in figure 2 ,
the protein levels of pgc-1 , tfam , and pparg were lower in the model group in comparison with the control group ( figure 2b2d ) , but nitroglycerin - induced reduction in the expression of pgc-1 , tfam , and pparg was inhibited by valproate .
the level of atp in the spinal trigeminal nucleus was measured in the nitroglycerin - induced trigeminovascular activation rat models with or without valproate treatment .
the atp level was lowered in the model group , indicating the dysfunction of mitochondrial energy metabolism .
compared with the model group , a low dose of valproate elevated the atp level , which was further increased by a high dose of valproate ( figure 3a ) .
these results suggest that nitroglycerin - induced dysfunction of mitochondrial energy metabolism in the spinal trigeminal nucleus is alleviated by valproate .
the mitochondrial membrane potential is crucial for energy metabolism in the mitochondria . in this study ,
as shown in figure 3b , almost all the jc-1 probes in the control group showed red fluorescence , which indicated a high mitochondrial membrane potential .
in contrast , the jc-1 probes in the model group showed green fluorescence , indicating a low mitochondrial membrane potential in the nitroglycerin - induced trigeminovascular activation rat model .
after treatment with a low dose of valproate , the jc-1 probes showed more red fluorescence and less green fluorescence , while the jc-1 probes in the model+vp - h group showed almost full red fluorescence ( figure 3b ) .
these results indicate that valproate treatment maintains the mitochondrial membrane potential in nitroglycerin - induced trigeminovascular activation .
consistent with the observations by fluorescence microscopy , the mitochondrial membrane potential in the model group was significantly decreased compared with the control group
. however , treatment with valproate prevented nitroglycerin - induced reduction of mitochondrial membrane potential ( figure 3c , 3d ) .
these results demonstrate that valproate preserves the mitochondrial membrane potential in the spinal trigeminal nucleus during nitroglycerin - induced trigeminovascular activation .
bax and bcl-2 have close relationships with the opening of the mitochondrial permeability transition pore . here , the protein levels of bax and bcl-2 were detected by western blot . in the model group ,
the protein level of bax was higher than that in the control group ( figure 4a ) , and the protein level of bcl-2 was lower than that in the control group ( figure 4b ) .
in contrast , treatment with valproate decreased the level of bax and increased the level of bcl-2 in the nitroglycerin - treated rats ( figure 4a , 4b ) .
these results provide molecular evidence for the hypothesis that valproate modulates the mitochondria energy metabolism in subjects with migraine .
as shown in figure 4c , the cytochrome c oxidase activity in the model group was decreased significantly as compared with the control group .
compared with the model group , valproate dose - dependently increased the activity of cytochrome c oxidase in the nitroglycerin - treated rats ( figure 4c ) .
consistent with the disrupted mitochondrial energy metabolism in nitroglycerin - treated rats , a significantly increased ros level was observed in the model group ( figure 4d ) .
treatment with valproate significantly attenuated nitroglycerin - induced ros elevation in the spinal trigeminal nucleus .
in the present study , we evaluated the effect of valproate on migraine in a rat model of nitroglycerin - induced trigeminovascular activation , and further investigated the underlying mechanism .
valproate decreased the number of scratching behaviors and reduced the changes in 5-ht and no levels in our rat model of nitroglycerin - induced trigeminovascular activation .
moreover , the mitochondrial biogenesis and energy metabolism , which was disrupted in nitroglycerin - treated rats , was stabilized by valproate .
these results prompted us to hypothesize that valproate may attenuate migraine by preserving mitochondrial function .
consistently , valproate has been reported to show a remission effect on migraine , and intravenous injection with valproate can rapidly abort headaches .
similarly , other antiepileptic drugs , such as topiramate and divalproex , have also been demonstrated to be effective prophylactic treatments of migraine .
mitochondria are crucial to mammalian cells because they are the major sites of atp production , and are also related to apoptosis regulation and ca homeostasis .
dysfunction of mitochondria can lead to a variety of disorders , especially in organs consuming large amounts of energy .
n - acetylaspartate ( naa ) , which is produced exclusively in neuronal mitochondria , is reported to be reduced in migraine patients .
recent reports also show that abnormal mitochondrial function leads to intracellular ca penetration , excessive ros production , and oxidative phosphorylation deficiency , which may ultimately cause energy failure in cells , and trigger migraine . in the present study
, we showed that the mitochondrial biogenesis and the mitochondrial energy metabolism were impaired in a rat model of nitroglycerin - induced trigeminovascular activation .
our results supportive the hypothesis that mitochondrial dysfunction is associated with the pathophysiology of migraine . in our study , the mtdna copy number was decreased in the nitroglycerin - induced trigeminovascular activation model , whereas treatment with valproate preserved the mtdna copy number in the spinal trigeminal nucleus .
the protein levels of pgc-1 , tfam , and pparg , which are closely associated with the biogenesis of mitochondria , were reduced in the rat model of nitroglycerin - induced trigeminovascular activation and were maintained by valproate .
the changes in these 3 proteins provide additional evidence for our hypothesis that valproate prevents dysregulation of mitochondrial biogenesis .
showed that valproate had positive effects on the maintenance of mtdna and mitochondrial biogenesis in polg - deficient fibroblasts .
moreover , valproate also plays an anti - oxidative role and alleviates neuronal damages caused by migraine attacks through nf-b .
the mitochondrial energy metabolism was disrupted in a rat model of nitroglycerin - induced trigeminovascular activation , and was rescued by valproate .
these results suggest that the effect of valproate on migraine may be associated with its action on mitochondrial function .
. showed that long - term treatment with valproate enhances mitochondrial function and protects against methamphetamine - induced mitochondrial damage .
however , komulainen et al . reported that treatment with valproate inhibits mitochondrial respiration and leads to mitochondrial dysfunction , oxidative stress , and increased cell death of hepg2 cells , which was assumed to be attributed to the hepatotoxicity of valproate .
interestingly , bachmann et al . also showed that a low dose of valproate enhances cellular respiration , whereas a high dose of valproate inhibits the respiration rate .
thus , the  contradictory  results from bachmann s and komulainen s work may be due to the different doses used . therefore , further investigations are needed to address whether valproate has hepatotoxicity during the treatment of migraine , as well as to determine the optimal dosage of valproate for the treatment of migraine .
the ratio of bax and bcl-2 is very important to the opening of the mitochondrial permeability transition pore , which also leads to the release of cytochrome c and the generation of ros .
migraine attack triggers oxidative stress , resulting in increased production of ros . in our study , nitroglycerin - induced mitochondrial dysfunction was associated with abnormal levels of bax , bcl-2 , cytochrome c oxidase , and ros , and these changes were diminished by valproate treatment .
as bax , bcl-2 , and ros are closely related to cell apoptosis , the abnormal levels of bax , bcl-2 , and ros in our study suggest that migraine may also have a relationship with apoptosis of neurocytes , but this hypothesis needs to be verified in future studies .
the protective effect of valproate against nitroglycerin - induced trigeminovascular activation was associated with valproate - mediated protection of mitochondrial biogenesis and function .
our study suggests that valproate may attenuate migraine through the maintenance of mitochondrial function , and provides a possible mechanism underlying the effect of valproate on migraine .
 backgroundmigraine is a chronic disease that interferes with life quality and work productivity . 
 valproate shows protective effects against migraine , yet the underlying mechanisms are unclear . this study aimed to evaluate the potential effect of valproate on migraine using a rat model of nitroglycerin - induced trigeminovascular activation , as well as to explore the underlying mechanism.material/methodsintraperitoneal injection of nitroglycerin 
 was conducted to induce trigeminovascular activation in rats . to explore the protective effect of valproate , a low dose ( 100 mg / kg ) or a high dose ( 200 mg / kg ) of valproate was intraperitoneally injected into rats , and then the levels of 5-hydroxytryptamine and nitric oxide in the peripheral blood were examined . 
 the mtdna copy number and the protein levels of peroxisome proliferator - activated receptor- coactivator 1 , mitochondrial transcription factor a , and peroxisome proliferator - activated receptor- in the spinal trigeminal nucleus were detected to evaluate the biogenesis of mitochondria . 
 the mitochondrial energy metabolism was determined by the mitochondrial membrane potential and the levels of adenosine triphosphate , cytochrome c oxidase , and reactive oxygen species.resultsvalproate attenuated nitroglycerin - induced trigeminovascular activation in rats , with reduced scratching behavior and restored 5-hydroxytryptamine and nitric oxide levels . moreover , 
 the mitochondrial energy metabolism and the biogenesis of mitochondria were preserved by valproate in nitroglycerin - treated rats.conclusionsthe protective effect of valproate against migraine may be achieved through the modulation of mitochondrial biogenesis and function . 
 our study provides evidence for the potential use of valproate in the treatment of migraine . 
Background
Material and Methods
Animals
Measurement of mtDNA copy number
Western blot analysis
Determination of adenosine triphosphate (ATP) level
Cytochrome C oxidase activity assay
Measurement of reactive oxygen species (ROS)
Determination of mitochondrial membrane potential
Statistical analysis
Results
Valproate attenuated nitroglycerin-induced trigeminovascular activation
Valproate modulated mitochondrial biogenesis
Mitochondrial energy metabolism was preserved by valproate
Discussion
Conclusions