Optical imaging of dendritic spikes in apical oblique dendrites of layer 5 pyramidal neurons

Abstract

Dendrites have active properties capable of generating dendritic spikes that could boost the impact of distal synaptic inputs. The strong passive filtering of the membrane and generation of local dendritic spikes enable different dendritic regions to function as independent computational compartments. While there is a wealth of information about cortical processing in apical tuft and basal dendrites, the functional role of apical oblique dendrites of layer 5 pyramidal neurons (L5PNs) are less understood. In this thesis, I aim to understand to functional role of thin apical oblique dendrites of L5PNs in the cortex. Using a previously published multi-compartment model of a L5PN, I first investigated the excitability of apical oblique dendrites and the extent of action potential (AP) back-propagation. In the model, I found that a 2-AP train at f > 35 Hz elicited an oblique branch spike in certain dendrites. The spike is mediated by activation of voltage-gated sodium and voltage-gated calcium channels. In addition, oblique branch spikes manifest as after-depolarizing potentials (ADP) at the soma. I then experimentally verified the generation of spikes by imaging the dendritic activity of oblique branches of L5PNs from the somatosensory cortex in vitro. I used our custom-built two-photon (2P) holographic microscope to perform functional calcium imaging on thin oblique branches of L5PNs in vitro. Oblique branch spikes are evoked at a critical frequency of fc = 57+/-5 Hz (from calcium imaging) and fc = 72+/-4 Hz (from ADP measurements) of a 2-AP train. Generation of spikes in oblique dendrites could establish their role as independent computational compartments that could boost coincident synaptic inputs. To further improve optical recording along dendrites, I proposed novel optical recording techniques to enhance the signal-to-noise ratio (SNRs) of the detected signals. A 4-fold improvement in the SNR was obtained with temporal gating for multi-site holographic calcium imaging, while a 6-fold increase in SNR was obtained for voltage imaging when using scattered photons to excite voltage indicators. These two novel SNR enhancement techniques can facilitate experiments that require imaging of activity in thin dendrites. Show more