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Brain-wide functional organization of the hippocampus along the dorsoventral axis: an optogenetic fMRI study
Russell W. Chan1,2,3, Eddie C. Wong1,2, Alex T. L. Leong1,2, Xunda Wang1,2, Celia M. Dong1,2, Karim E. Hallaoui1,2, and Ed X. Wu1,2

1Laboratory of Biomedical Imaging and Signal Processing, The University of Hong Kong, Hong Kong, China, 2Department of Electrical and Electronic Engineering, The University of Hong Kong, Hong Kong, China, 3Neurology and Neurological Sciences, Stanford University, Stanford, CA, United States

Synopsis

Hippocampus plays a prominent role in central nervous system functions. It receives convergent projections and sends reciprocal divergent projections, forming an interactive cortico-hippocampal-cortical network. However, the precise brain-wide functional organization of different hippocampal activities along dorsoventral axis remains unknown. Using optogenetic fMRI, we revealed that functional organization of low frequency hippocampal activities along dorsoventral axis exhibits a gradual change from regions mainly involved in cognition and sensory processing to regions also involved in motor control and anxiety-related behavior. Additionally, hippocampal activities generally transit from long-range propagation to downstream cortical/subcortical regions to local intra-hippocampal propagation with increasing frequencies.

Purpose

Hippocampus (HP) plays a prominent role in central nervous system functions such as learning1-3, memory4,5, sensory processing6, spatial navigation7,8, motor control9,10, emotions11,12 and stress13. The HP, including the dentate gyrus (DG), can evoke large-scale influences on cortical and subcortical activities, since HP receives convergent projections from sensory and limbic regions before sending reciprocal divergent projections to form a highly interactive cortico-hippocampal-cortical network and subcortico-hippocampal-subcortical network14,15. At present, the precise functional role of these projections remains a topic of much debate. Previous anatomical and electrophysiological studies revealed that hippocampal connectivities with entorhinal cortex14,16 and amygdala17 show dorsoventral topographical gradients, indicating multiple functional gradients along the dorsoventral axis18,19. Meanwhile, gene expression studies20,21 also demonstrated multiple genetic domains along the axis, signifying discrete functional domains18,19. Our recent fMRI study demonstrated that low frequency (1 Hz), but not high frequency (40 Hz), optogenetic stimulation of excitatory neurons in anterior dorsal DG (adDG) evoked robust cortical and subcortical brain-wide fMRI responses22. However, how hippocampal activities are functionally and dynamically organized along the dorsoventral axis remains largely unknown. Since dentate gyrus (DG) primarily receives cortical and subcortical terminal projections and acts as a bridge for cortico-hippocampal-cortical and subcortico-hippocampal-subcortical communication, we examined how different frequencies of hippocampal activity initiated in DG along dorsoventral axis contribute to brain-wide functional organization by combining optogenetic23, cell-specific stimulation of CaMKIIα-expressing excitatory neurons and large-scale fMRI detection22,24,25.

Materials and Methods

Animal preparation: As illustrated in Fig. 1, AAV5-CaMKIIα::ChR2(H134R)-mCherry was injected to posterior dorsal DG (pdDG) (n=5) or ventral DG (vDG) (n=6) of adult male SD rats. After 4-weeks, an optical fiber was implanted to deliver optogenetic stimulation.

Optogenetic stimulation: Blue light (473nm; 40mW/mm2; 1Hz, 5Hz, 10Hz, 20Hz and 40Hz) was used to determine frequency-dependent spatiotemporal characteristics of evoked pdDG and vDG responses. Two-block design of 20s light-on and 140s light-off was applied.

Optogenetic fMRI acquisition and analysis: fMRI was performed on a Bruker 7T scanner using GE-EPI (FOV=32×32mm2, matrix=64×64, α=56°, TE/TR=20/1000ms). Standard preprocessing and GLM analysis were applied to identify BOLD responses (P<0.001). BOLD signal profiles were then extracted from anatomically defined regions.

Results

Anatomical MRI scans and histology conīŦrmed viral injection, fiber implantation, and ChR2-mCherry expression in pdDG and vDG (Fig. 1). pdDG 1Hz stimulation evoked positive responses in ipsilateral dorsal HP (dHP) and ventral HP (vHP), bilateral primary visual cortex (V1), primary auditory cortex (A1) and cingulate cortex (Cg), while 5Hz and 10Hz evoked robust activations in bilateral vHP, dHP, entorhinal cortex (Ent), V1, A1, primary somatosensory cortex (S1), Cg, retrosplenial cortex (RS), primary motor cortex (M1), caudate putamen (CPu) and amygdala (AM). We also stimulated pdDG at 20Hz and 40Hz. Strong responses were observed in bilateral hippocampus (Fig. 2).

vDG 5Hz and 10Hz stimulation evoked robust positive responses in bilateral vHP, dHP, Ent, V1, A1, S1, Cg, RS, M1, CPu, AM and hypothalamus (hTh), which are regions related to learning, memory, sensory processing, motor control, emotions and stress. These cortical and subcortical responses decreased at 20Hz and 40Hz, while strong responses retained in vHP and dHP (Fig. 3). Note that no evoked responses were observed in naïve animal (data not shown), indicating that the observed responses were a direct result of optogenetic stimulation, not the heat-induced artifacts26.

Discussions and Conclusion

In this study, stimulations of pdDG (Fig. 2) revealed a shift from regions actively involved in only cognition and sensory processing (1Hz) to regions also associated with motor control and anxiety-related behavior (5-10Hz), and then to local hippocampal regions (20-40Hz). For vDG stimulations (Fig. 3), spatiotemporal response properties shifted from regions actively involved in cognition, sensory processing, motor control and anxiety-related behavior (5-10Hz) to local hippocampal regions (5-40Hz). Our recent study demonstrated that both BOLD response upon anterior dorsal dentate gyrus (adDG) stimulations shifted from regions actively involved in cognition and sensory processing (1Hz) to local hippocampal regions (40Hz)22.

Together, our optogenetic fMRI findings here clearly revealed that, along the dorsoventral axis (i.e., from adDG, pdDG to vDG), the downstream targets of the low frequency hippocampal activities (1-10Hz) shift from regions only actively involved in cognition and sensory processing to regions also associated with motor control and anxiety-related behavior. Furthermore, with increasing frequency, hippocampal activities generally transit from long-range propagation to downstream cortical/subcortical regions to local intra-hippocampal propagation. These results contribute to our present understanding of hippocampal activities are functionally and dynamically organized along the dorsoventral axis.

Acknowledgements

This work was supported by the Hong Kong Research Grant Council (Grants C7048-16G and HKU17103015 to E.X.W.).

References

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Figures

Figure 1. Experimental setup for optogenetic stimulation and histological characterization of CaMKIIα::ChR2 viral expression in (A) posterior dorsal dentate gyrus (pdDG) and (B) ventral dentate gyrus (vDG). Schematic (Left) and T2-weighted anatomical images (Middle) show the viral injection and fiber implantation sites, respectively. Histology images (Right, green box) show respective ChR2 expression in pdDG and vDG.

Figure 2. (A) Averaged activation maps of optogenetic stimulation in pdDG. Positive BOLD responses detected in ipsilateral dHP and vHP, and bilateral V1, A1 and Cg during 1Hz stimulation (n=5; t>3.1, corresponding to P<0.001). For 5Hz stimulation, robust activations observed in bilateral vHP, dHP, Ent, V1, A1, S1, Cg, RS, M1, CPu and AM (n=5; t > 3.1, corresponding to P < 0.001). (B) ROIs definitions to extract BOLD signal profiles. Abbreviations: Please refer to main text. (C) The respective BOLD signal profiles extracted from the ROIs.

Figure 3. (A) Averaged activation maps of optogenetic stimulation in vDG. Robust positive BOLD responses detected in bilateral vHP, dHP, Ent, V1, A1, S1, Cg, RS, M1, CPu, AM and hTh during 5Hz stimulation (n=6; t>3.1, corresponding to P<0.001). (B) ROIs defined by rat brain atlas to extract BOLD signal profiles. Abbreviations: Please refer to main text. (C) The respective BOLD signal profiles extracted from the ROIs. Error bars indicate ±SEM.

Proc. Intl. Soc. Mag. Reson. Med. 26 (2018)
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