Methods used in the Febo laboratory
Our laboratory seeks to determine mechanistic links between in vivo whole brain activity and affective behavioral responses in mouse models of Alzheimer’s disease. However, due to the restricted capacity for movement of mice inside the MRI scanner, it is difficult to achieve this goal. Although there exist approaches that can measure localized activity of neuronal populations while animals display specific behavioral patterns, these have limited capacity to capture network level activity. One way to overcome this limitation is to combine optogenetic methods with fMRI and behavior.
The above figure illustrates this approach (Tan et al, Journal of Neuroscience, 2019, 39(17):3249-3263)). Below are several of the methods we have available in our laboratory (MBI L1-135 and L4-129). In addition to these MRI and behavioral methods, our laboratory is supplied for histological assessments in collaboration with the UF pathology core and the UF Interdisciplinary Center for Biotechnology Research (ICBR). We also have been recently started a collaboration with LifeCanvas Technologies in Cambridge MA to conduct light sheet microscopy to evaluate amyloid plaque and inhibitory neuron distribution in cleared whole brain preparations.
Functional magnetic resonance imaging
We use AMRIS facility MRI scanners that are housed in the McKnight Brain Institute ground floor. These include 11.1 Tesla and a 7 Tesla scanners. We also greatly benefit from the availability of custom in-house and Bruker radiofrequency systems, all maintained by outstanding and supportive AMRIS staff.
The fMRI method relies on the BOLD weighting (T2 or T2*) of echo planar images to detect with enough sensitivity MR signals coupled to the ratio in transient concentrations of oxy-to-deoxyhemoglobin (diamagnetic-to-paramagnetic effects on proton relaxation times, respectively). The BOLD signal increases as this ratio increases and it falls to levels as this ratio recovers to a baseline state. These neurometabolic signals are linked to changes in synaptic activity in neocortical somatodendritic inputs. Indeed, BOLD signals are an ‘indirect’ or surrogate measure of neuronal activity.
The images have considerably low resolution relative to other invasive approaches. The signal intensity or contrast relative to a baseline state of a single T2/T2* weighted fMRI voxel in cortex emerges from the activity of tens of thousands of neurons (~50,000 according to some estimates). Thus, the fMRI signal is a ‘net’ signal representing the summation of intravoxel biological signals from a cooperative neuronal-astrocytic-microvasculature system in neocortex. Regional brain changes in BOLD are thus a weighted intervoxel sum of cortical substrates that are involved in active states and also in the processing of ‘neural activity’ under resting conditions. Thus, fMRI represents largescale activity that be used to derive features from macroscopic brain networks. One of the challenges is to ‘back-reconstruct’ the synaptic activity of neocortical microcircuits that give rise to the BOLD signal. The combined use of fMRI along with lightsheet microscopy could be a platform to partly achieve this.
Diffusion weighted imaging
Will soon be updated!
Lightsheet microscopy
3-dimensional whole brain lightsheet microscopic imaging (LSI) in mouse brain tissue that is cleared and triple labeled (e.g., below are plaques and PV+ cells, along with a general cell nuclear stain like propidium iodide) is a new approach we are using to explore functional and tissue histological changes with high precision. Both fMRI and LSI brains are aligned to the same template to enable multiple assessments in the same mouse. The mouse brain template was kindly provided by Dr. G. Allan Johnson, Center for In Vivo Microscopy, Duke).
Behavioral testing in mice
Locomotor activity
We have 4 automated activity cages from Accuscan Instruments in MBI L4-129. These can be used for mouse and rat. In mice the cages may be used in novel open field tests for anxiety like behaviors.
Fear conditioning
We have 3 Med-Associates Operant Cage Systems for Fear Conditioning in Mice. These can be used for classical and contextual fear conditioning. Cages are located in Febo lab L1-135A.
Sociability/social recognition
Two custom plastic test cages for sociability tests are located in L1-135A in the Febo lab. These can also be used in test for novel object recognition.
Sucrose preference testing
We use leak proof mouse cage bottles from Braintree Scientific for home cage sucrose consumption tests.
Tail suspension test
We have 4 Med-Associates Tail Suspension test boxes with control computer running Med-PC and Tail Suspension Software. Cages are located in Febo lab L1-135B.
T-maze test
We have a modular T-maze from Med-Associates. the cage is fully automated with sliding doors dividing compartments and infrared detection beams for each arm and for head entry ports for rewards are controlled by Med PC software. Cage is located in Febo lab L1-135A.