| The brain is the basis of high-level neural activity,which dominates all activities in the body.Brain damage or the abnormalities of its structural and functional can affect the normal functioning of the body.Understanding the anatomical structure and physiological functions of the brain is of great significance for the treatment of brain injury diseases.The secondary motor cortex(MOs)is located in the anterior lateral area of the brain cortex in rodents.Recent studies have provided evidence for potential roles of the MOs in cognitive and executive processes,such as decision making,goal-directed actions,skill learning,and spatial memory.Impairment or inactivation of the MOs results in neglect of the contralateral space in movement and motor order learning impairment.To data,most knowledge of the MOs focus on one direct pathway from the MOs to its connected regions,resulting in a lack of knowledge of the overall projections of the MOs and severely hindering the comprehensive understanding of how the MOs is wired to process information at a global scale.To address this issue,we sparse labeled pyramidal neurons nearby or in the MOs using a Cre-dependent adeno-associated virus(AAV).In whole brian imaging,comparing the data quality obtained by optical clearing and fMOST,the fMOST was selected for the acquisition of single neuron morphology of MOs.So far,the protocol was established.Labeling the neuron with recombinant adeno-associated virus,acquiring the whole brain data with fMOST,and manually reconstructing the anatomical morphology of neurons.According the protocol,we acquired an uninterrupted whole-brain 3D dataset at a voxel resolution of 0.2 × 0.2 × 1 ?m with a whole-brain fluorescence imaging system(fMOST).Based on the 3D dataset,we successfully reconstructed the complete morphologies of 38 brightly labeled neurons nearby or in the MOs,which consisted of 8 neurons in layer5 of MOs,17 neurons in layer2/3 of MOs,6 neurons in layer2/3 of PL,and 7 neurons in layer2/3 of ORBm.Furthermore,we analyzed the projection patterns and projection strengths of these neurons at a single-neuron level based on several parameters,including the projection areas,the fiber length,the total number of terminal tips,and the total number of branches.The main findings are as follows:(1)The axonal lengthes of layer 2/3 neurons range from 60 mm to 80 mm.Most of the neurons in layer5 have an axon length of more than 200 mm,the longest of which is 318.43 mm,which is the longest of the reconstructed neuons to data.(2)The projection patterns of the neurons in the MOs and its surrounding areas,such as the prelimbic cortex(PL)and the medial orbital cortex(ORBm),are different.However,the axons and dendrites of the layer2/3 neurons are similar in length,suggesting that neurons in different regions have similar inputs and outputs but participates in different brain functions.(3)The anxonal length of the commissural projection neurons(CPN)in layer5 of the MOs is 3 to 4 times that of the layer2/3 neurons.All these CPNs have a dense axonal projection in the striatum region,which may be related to serial order performance.Although the neurons in layer5 all have strong projectiont in the striatum,the projection pattern in other cerebral cortex are quite different.In a word,there is no two neurons have same projection patterns in this study.In summary,we obtained uninterrupted complete morphologies of 38 IT neurons nearby or in the MOs based on our whole-brain 3D dataset and analyzed the projections of these neurons.To the best of our knowledge,this study is the first to show completed morphologies of individual reconstructed neurons in the MOs.Our results reveal the diversity of the projection patterns for neurons in the same brain region and the complexity of the axonal projections in a single brain region.Our results will be helpful for further understanding the wiring diagram of the MOs at the level of a single neuron and lay a solid foundation for exploring the behavioral functions of the MOs. |
PDF Full Text Request