| PartⅠ:Expression of different isoforms of Homer in different encephalic regions of SHR/WKYObjective:To investigate the effect of Homer on pathogenesis of Attention Deficit/Hyperactivity Disorder(ADHD).Method:(1)To characterize behavioural alterations,we studied locomotor activity of juvenile Spontaneously hypertensive rat(SHR)/Wistar Kyoto(WKY)by using open-field environment test.(2)The total RNA and protein were extracted from prefrontal cortex,hippocampus,nucleus accumbens and corpus striatum of SHR and WKY respectively.Both RT-PCR and western blotting technique were used to examine the expression of different isoforms of Homer(Homer la,Homer 1b/c,Homer 2a/b)in different encephalic regions of SHR and WKY.Results:(1)Ambulatory and rearing activities in the open-field environment were significantly higher in SHR than in their controls(t=8.34,p＜0.01;t=4.43,p＜0.01).(2) By RT-PCR detection,it was found that the expressive levels of Homer 1a gene were significantly lower in prefrontal cortex and hippocampus of SHR than those in WKY(t=2.56,p＜0.05;t=2.95,p＜0.05),and the levels of Homer 2a/b genewere significantly lower in prefrontal cortex, hippocampus and nucleus accumbens of SHR than those in WKY(t=5.85, p＜0.01;t=2.66,p＜0.05,t=2.66,p＜0.05).There was no difference between the expressive levels of Homer 1a gene in nucleus accumbens and corpus striatum of SHR and WKY,and there was also no difference between the expressive levels of Homer1b/c gene in prefrontal cortex, hippocampus,nucleus accumbens and corpus striatum of SHR and WKY, and the same as the expressive levels of Homer 2a/b gene in corpus striatum of SHR and WKY.(3)By western blotting,it was found that the expressive levels of Homer 1a gene were significantly lower in prefrontal cortex and hippocampus of SHR than those in WKY(t=2.76,p＜0.05; t=4.31,p＜0.01),and the expressive levels of Homer 2a/b gene were also significantly lower in prefrontal cortex and hippocampus of SHR than those in WKY(t=3.40,p＜0.05;t=3.27,p＜0.01).Conclusion:(1)Our findings reveal that juvenile SHR manifest behaviours resembling attention deficit/hyperactivity disorder(ADHD)in hyperactivity.(2) Homer 1a,Homer 2a/b may play an important role in pathogenesis of ADHD. PartⅡ:Effects of methylphenidate on behavior and expression of Homer and some related genes of SHRObjective:To investigate the effects of methylphenidate(MPH)on behaviors of SHR and to explore the effects of Homer on pharmacology of methylphenidate in curing ADHD.Method:(1)We studied the effects of acute(2mg/kg i.g.)or chronic(2mg/kg i.g.bid for 14days) administration of methylphenidate on locomotor activity and attention of the experimental group(MPH group)and the control group(Ctl group) by using open-field test and Làt maze,as well as on learning and memory by using Morris water maze.All the behavior in the tests were monitored by a CCD video camera and analyzed off-line.(2)The total RNA and protein were extracted from prefrontal cortex and hippocampus of SHR and WKY respectively.Both RT-PCR technique and western blotting technique were used to detect the expressive levels of Homer 1a,Homer 2a/b and some related gene(eg.mGluRI,IP3R,Shank)in prefrontal cortex and hippocampus of the MPH group and the Ctl group.Results:(1) Before acute administration,there was no difference in ambulatory and rearing activities in the open-field environment between the experimental group and the control,as well as the ambulatory and rearing activities and the frequency of leanings against the glass wall in a Làt maze before chronic administration.It was found that the ambulatory and rearing activities and the frequency of leanings against the glass wall in a Làt maze after acute administration all revealed decreasing trends in the MPH group compared to the Ctl group.However,the ambulatory and rearing activities and the frequency of leanings against the glass wall were significantly lower in the MPH group than the control(t=3.63,p＜0.05;t =3.24,p＜0.05;t=3.98,p＜0.05)after chronic administration.Meanwhile, in Morris water maze,the total distances swimmed were significantly lower in MPH group than the control during the second and third days(t =2.36,p＜0.05;t=2.30,p＜0.05)rather than the first day,and the same as the latencies(t=2.76,p＜0.05;t=2.51,p＜0.05).(2)By RT-PCR detection,it was found that the expressive levels of Homer 1a gene rather than Homer 2a/b gene or other related genes were significantly higher in prefrontal cortex and hippocampus of the experimental group and the control(t=2.41,p＜0.05;t=2.68,p＜0.05)after acute administration. However,it was found that the expressive levels of Homer 1a,Homer 2a/b,Shank 1(t=2.82,p＜0.05;t=2.73,p＜0.05;t=3.34,p＜0.05)rather than mGluR 1,mGluR 5,IP3R 1,IP3R 2,IP3R 3,Shank 2,Shank 3 were siginificantly higher in prefrontal cortex of the MPH group than the control,and the same as in hippocampus(t=3.12,p＜0.05;t=2.36, p＜0.05;t=2.74,p＜0.05).(3)By western blotting detection,it was found that the expressive levels of Homer 1a rather than Homer 2a/b were significantly higher in prefrontal cortex and hippocampus of the experimental group and the control(t=3.21,p＜0.05;t=4.07,p＜0.05) after acute administration.However,it was found that expressions of Homer 1a were significantly higher in prefrontal cortex,hippocampus of the experimental group than the control(t=4.41,p＜0.05;t=3.04,p＜0.05), and the same as Homer 2a/b(t=2.62,p＜0.05;t=2.87,p＜0.05)after chronic administration.Conclusion:(1)Our findings revealed that juvenile SHR by acute or chronic administration of oral methylphenidate manifest the coincidence behaviours resembling ADHD patients after curing in clinical.(2)Both Homer 1a and Homer 2a/b may play extremely important roles not only in pathogenesis of ADHD but also in pharmacology of methylphenidate. |
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