| BackgroundTourette Syndrome(TS),a commonly diagnosed neurological pediatric ailment,features multiple short and stereotypical motor and phonic tics.The main clinical manifestations are head,face,abdomen and limbs involuntary twitching,voice of larynx.It is estimated that 0.3%-0.9%of children worldwide have TS.Average age at TS onset is 6-7 years,and 93%of TS patients develop symptoms before the age of 10 years.Symptoms vary from mild to severe,and one half to two thirds of the affected children are virtually symptom free or experience a substantial decrease in tics before age 18.TS affects males more frequently than females and is often associated with attention deficit hyperactivity disorder(AD HD)and obsessive-compulsive disorder(OCD).There is frequently an urge that precedes the tic,sometimes in the form of a specific sensory feeling(a sensory tic).Patients state that they perform the tic in order to reduce the urge,although shortly after the tic manifests,the urge recurs.TS can interfere and damage children’s cognitive function and development,and affect their social adaptability.At present,medicinal treatment is the main therapeutic option for reducing tics in TS patients.Indeed,the typical antipsychotics haloperidol and pimozide effectively reduce tic frequency.However,undesirable effects,including extrapyramidal symptoms(EPS)and tardive dyskinesia(TD),occur after haloperidol administration.Other treatments include habit-reversal cognitive behavioral therapy,and localized intramuscular administration of botulinum toxin,with serious,refractory cases responding to deep brain stimulation surgery(DBS).However,none of these methods can completely cure the disease.Traditional Chinese Medicine(TCM)is a practice that has been used for thousands of years due to its apparent effectiveness and infrequent side effects.Ningdong granule(NDG),a compound preparation used in TCM,is applied to alleviate tics,for which it has been indicated to be effective.As the major active component of Gastrodia elata,a Chinese herb,gastrodin(Gas)has been widely used since its first description in 1978,with a wide range of beneficial effects on headache,migraine,dizziness,infantile convulsions,and tetany.3,3’-Iminodipropionitrile(IDPN)has frequently been used to develop animal models of tics,as it can cause persistent behavior syndromes that include head shaking,random circling,hyperactivity and increased acoustic startle responses.These stereotyped behaviors are similar to the symptoms of TS.In the present study,the focus was on the DA and 5-HT systems in order to evaluate the anti-tic function of NDG and Gas in an animal model.Micro-positron emission tomography(PET)was used to evaluate the binding of D2Rs,DA transporters(DATs),5-HTIA receptors(5-HT2ARs)and 5-HT transporters(SERTs)in brain regions of interest(ROIs)。Our aim was to explore the effects of NDG and Gas on the DA and 5-HT system of TS rat models,and clarify the molecular mechanism of NDG and Gas in the treatment of TS.Objective1.To observe the effects of Ningdong granule and Gas on TS rat models.2.To observe the effects of Ningdong granule and Gas on the contents of DA receptor,DA transporter,5-HT receptor and 5-HT transporter in brain of TS rats.To investigate the mechanism of Ningdong granule and Gas in the treatment of TS.Methods1.Experimental animalsThirty Wistar rats(male,150 ± 10 g),were maintained in airconditioned animal rooms under a 12-h light/dark cycle at 22±2℃and 50±10%humidity.The rats were allowed drinking water and rodent chow at will,and fed for 1 week to adapt to the new environment.Then the animals were randomized to the control(n=6)and four experimental(IDPN+NS,IDPN+Hal,IDPN+NDG,and IDPN+Gas)groups(n=6/group).The control group received intraperitoneal injections(i.p.,5 ml/kg)of normal saline(NS,0.9%)daily,while the experimental groups were administered daily IDPN injections(150 mg/kg,i.p.)for 7 consecutive days.Upon TS induction for 7 days,the experimental rats were intragastrically treated daily for eight weeks with either normal saline(0.9%)at 10 ml/kg(IDPN+NS and control groups),Hal at 1.0 mg/kg(IDPN+Hal group),NDG at 22 g/kg(IDPN+NDG group),or Gas at 20 mg/kg(IDPN+Gas group).2.Micro-PETBefore micro-PET,the animals were anesthetized with isoflurane and administered by tail vein injection with approximately 150-250 ul(250± 50 uCi)of 18F-fallypride(mostly binding to D2Rs),18F-FECNT(interacts with DATs),18F-altanserin(interacts 5-HT2ARs)or 18F-FPBM(interacts with SERTs).The rats were allowed to uptake the radiotracer for ten minutes.Then,each rat was positioned prone on a platform placed in the micro-PET equipment and scanned for 10 min.Image reconstruction was performed interactively with an ordered subsets expectation maximization(OS-EM 3D)algorithm based on 128*128*159 image matrices,yielding pixel size and slice thickness of 0.77 mm and 0.78 mm,respectively.Anatomical regions of interest(ROIs),cerebral basal ganglia,were delineated using the PMOD software(Version3.7,USA),and tracer uptake was calculated by empirical formulae.To empty their bodies of other reagents,the rats underwent scans once every week.3.Research contents(1)The behavior changes of rats after medication measured by behavior score;(2)Analysis the variation of D2R,DAT,5-HT2AR,SERT in the brain of rats by Micro-PET.Results1.Behavioral evaluationIDPN caused abnormal,stereotypical behaviors in rats.After 8 weeks of treatment,these abnormal behaviors were significantly reduced in the NDG group;Gas group and Hal group,compared with the blank control group(P<0.05).There was no statistically significant difference in the behaviors of rats between NDG;Gas and Hal groups.2.Micro-PET imagingD2R.In the striatum,IDPN increased the level of D,R binding compared with that in the control group(P<0.05).Following treatment,NDG,Gas or Hal significantly reduced the upregulation in striatal D2R content(P<0.01).No significant difference was observed between the NDG,Gas and Hal groups.DAT.The DAT content in the striatum was higher in the IDPN group than in the control group(P<0.05).Following treatment with NDG and Gas,the DAT content in the striatum was increased compared with that in IDPN group(P<0.05).However,no significant difference was determined in the Hal group compared with the IDPN group.5-HT2AR.There was a slight increase in the 5-HT2AR content in the striata of TS model rats,though this was not significant when compared with the content in control rats.There were no differences in the 5-HT2AR content in other brain regions among the groups.After 8 weeks of NDG administration,the uptake ratio of 18F-altanserin was significantly increased in the striatum and cerebral cortex compared with IDPN group(P<0.05).Meanwhile,Gas was slightly increased the density of 5-HT2AR in the basal ganglia though this was not significant compared with that in control group.However,Hal failed to stimulate any changes in 5-HT2AR content(P)0.05).SERT.IDPN-treated rats exhibited no significant differences relative to the control group for all brain ROIs.Following treatment,NDG significantly reduced the SERT content in the nucleus accumbens,putamen,amygdaloid nucleus,striatum and cerebral cortex compared with that in IDPN group(P<0.01).Gas reduced the density of SERTs significantly in the basal ganglia compared to IDPN group(P<0.05).Hal reduced the SERT content in the nucleus accumbens and putamen compared to IDPN group(P<0.05).However,in other brain regions there were no significant differences between these groups.ConclusionIn conclusion,Ningdong Granule and Gastrodin can effectively alleviate the inflexible behavior of TS rats.The mechanism of NDG is to directly inhibit the excitability of DA system by decreasing the content of DA receptor and increasing the content of DAT,and indirectly inhibit the excitability of DA system and promote DA metabolism by increasing the content of 5-HT receptor and decreasing the content of SERT.Gas can relieve twitching mainly through the regulation of DA system,but has little effect on 5-HT system. |
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