Experimental Study On Behavioral Changes Induced By Low Intensity Focused Ultrasound Stimulation Of The Nucleus Accumbens In Mice

Objective:C57BL/6 common mice were used in this study,the nucleus accumbens of common mice was selected for ultrasound stimulation to seek the effectiveness of ultrasonic stimulation.Using low intensity focused ultrasound in mice for ultrasound stimulation,then doing test,including forced swimming,open field test,Morris water maze test,and transmission electron microscopy,enzyme-linked immuneosorbency assays.And exploring its possible mechanism of action,to provide new possibilities and experimental data for researching the mechanism of ultrasound neuromodulation,and to propose new methods for the treatment of diseases.Methods:1.Ultrasound stimulation:100 C57BL/6 mice were randomized into control and US group.The control group was given sham stimulation,and the US group was given low-intensity focused ultrasound to stimulate the NAc.Mice were anesthetized with 1.5%isoflurane via inhalation and the head shaved,and the mice were fixed on the brain stereotaxer.The location of NAc was found according to the brain map of the mice(with Bregma as the coordinate origin,X axis+0.05cm,Y axis+0.118cm).7 days of US(0.5MHz,50%duty cycle,1000 Hz pulse repetition frequency,10 min/day,twice a day)was delivered to NAc.The control group operated in the same way,but the ultrasound device was not activated.After the intervention,30 mice were randomly assigned to behavioral assessment(forced swimming,open field,and water maze),10 mice were assigned to electron microscopy(EM),and 10 mice were assigned to neurotransmitter test.2.Behavioral test:Tests were conducted immediately after the intervention.The forced swimming test assessed depression in mice by measuring the immobile time in the water.The open field test evaluated the anxiety level of mice by measuring the total distance travelled,the average speed travelled,the distance travelled of the central area,the time of the central area and the number of exits from the central area.In the water maze experiment,the learning and memory ability of mice was evaluated by taking the opposite quadrant of the target quadrant(the third quadrant)as the entry point,the avoidance latency time(the time taken for the mice to climb the platform),the number of crossing the platform,and the residence time in the target quadrant.3.Indicator detection:NAc was sampled immediately after the intervention.The ultrastructural changes of NAc neurons and the monoamine neurotransmitters Norepinephrine(NE),Dopamine(DA)and 5-hydroxytryptamine(5-hydroxytryptamine)were detected by transmission electron microscopy and ELISA.To explore the mechanism of neuroregulation by ultrasound.Results:1.Behavioral test results:Compared with the control group,the immobile time in the US group was significantly longer(P<0.001),the total distance travelled,the average speed travelled,the distance travelled of the central area,the time of the central area and the number of exits from the central area in the open field all decreased significantly(P<0.05,P<0.01),In the water maze experiment,the avoidance latency time was significantly longer,the number of crossing the platform was significantly reduced,and the residence time in the target quadrant was significantly shorter(P<0.05,P<0.001).2.Electron microscopy results:The ultrastructure of NAc neurons in the control group was basically normal.In the US group,the neurons were swelling,nerve fiber dissolution and necrosis,mitochondrial vacuolation,endoplasmic reticulum dilation,and intracytoplasmic organelles were reduced or disappeared.3.Measurement results of neurotransmitters:Compared with the control group,the contents of NAc monoamine neurotransmitters NE,DA and 5-HT were significantly decreased in the US group(P<0.0001).Conclusion:1.Low intensity focused ultrasound stimulation changed the excitability of normal mice to special environment,and reduced their ability to explore new environment,adapt to new environment,learning and memory ability,and showed symptoms similar to depression and anxiety.2.Low intensity focused ultrasound stimulation induced the morphological changes of NAc neurons and the decrease of monoamine transmitters,which may be the cause of depression and anxiety and the decrease of learning and memory ability in mice.