def run_volpy(fnames,
options=None,
do_motion_correction=True,
do_memory_mapping=True,
fr=400):
#pass # For compatibility between running under Spyder and the CLI
# %% Load demo movie and ROIs
file_dir = os.path.split(fnames)[0]
path_ROIs = [file for file in os.listdir(file_dir) if 'ROIs_gt' in file]
if len(path_ROIs) > 0:
path_ROIs = path_ROIs[0]
#path_ROIs = '/home/nel/NEL-LAB Dropbox/NEL/Datasets/voltage_lin/peyman_golshani/ROIs.hdf5'
#%% dataset dependent parameters
# dataset dependent parameters
fr = fr # sample rate of the movie
# motion correction parameters
pw_rigid = False # flag for pw-rigid motion correction
gSig_filt = (3, 3) # size of filter, in general gSig (see below),
# change this one if algorithm does not work
max_shifts = (5, 5) # maximum allowed rigid shift
strides = (
48, 48
) # start a new patch for pw-rigid motion correction every x pixels
overlaps = (24, 24
) # overlap between pathes (size of patch strides+overlaps)
max_deviation_rigid = 3 # maximum deviation allowed for patch with respect to rigid shifts
border_nan = 'copy'
opts_dict = {
'fnames': fnames,
'fr': fr,
'pw_rigid': pw_rigid,
'max_shifts': max_shifts,
'gSig_filt': gSig_filt,
'strides': strides,
'overlaps': overlaps,
'max_deviation_rigid': max_deviation_rigid,
'border_nan': border_nan
}
opts = volparams(params_dict=opts_dict)
# %% start a cluster for parallel processing
c, dview, n_processes = cm.cluster.setup_cluster(backend='local',
n_processes=None,
single_thread=False)
# %%% MOTION CORRECTION
# first we create a motion correction object with the specified parameters
mc = MotionCorrect(fnames, dview=dview, **opts.get_group('motion'))
# Run correction
do_motion_correction = do_motion_correction
if do_motion_correction:
mc.motion_correct(save_movie=True)
else:
mc_list = [
file for file in os.listdir(file_dir)
if (os.path.splitext(os.path.split(fnames)[-1])[0] in file
and '.mmap' in file)
]
mc.mmap_file = [os.path.join(file_dir, mc_list[0])]
print(f'reuse previously saved motion corrected file:{mc.mmap_file}')
# %% MEMORY MAPPING
do_memory_mapping = do_memory_mapping
if do_memory_mapping:
border_to_0 = 0 if mc.border_nan == 'copy' else mc.border_to_0
# you can include the boundaries of the FOV if you used the 'copy' option
# during motion correction, although be careful about the components near
# the boundaries
# memory map the file in order 'C'
fname_new = cm.save_memmap_join(
mc.mmap_file,
base_name='memmap_' +
os.path.splitext(os.path.split(fnames)[-1])[0],
add_to_mov=border_to_0,
dview=dview) # exclude border
else:
mmap_list = [
file for file in os.listdir(file_dir)
if ('memmap_' +
os.path.splitext(os.path.split(fnames)[-1])[0]) in file
]
fname_new = os.path.join(file_dir, mmap_list[0])
print(f'reuse previously saved memory mapping file:{fname_new}')
# %% SEGMENTATION
# create summary images
img = mean_image(mc.mmap_file[0], window=1000, dview=dview)
img = (img - np.mean(img)) / np.std(img)
gaussian_blur = False # Use gaussian blur when there is too much noise in the video
Cn = local_correlations_movie_offline(mc.mmap_file[0],
fr=fr,
window=fr * 4,
stride=fr * 4,
winSize_baseline=fr,
remove_baseline=True,
gaussian_blur=gaussian_blur,
dview=dview).max(axis=0)
img_corr = (Cn - np.mean(Cn)) / np.std(Cn)
summary_images = np.stack([img, img, img_corr], axis=0).astype(np.float32)
# ! save summary image, it is used in GUI
cm.movie(summary_images).save(fnames[:-5] + '_summary_images.tif')
#plt.imshow(summary_images[0])
#%% three methods for segmentation
methods_list = [
'manual_annotation', # manual annotation needs user to prepare annotated datasets same format as demo ROIs
'gui_annotation', # use gui to manually annotate neurons, but this is still under developing
'maskrcnn'
] # maskrcnn is a convolutional network trained for finding neurons using summary images
method = methods_list[0]
if method == 'manual_annotation':
#with h5py.File(path_ROIs, 'r') as fl:
# ROIs = fl['mov'][()]
ROIs = np.load(os.path.join(file_dir, path_ROIs))
elif method == 'gui_annotation':
# run volpy_gui file in the caiman/source_extraction/volpy folder
# load the summary images you have just saved
# save the ROIs to the video folder
path_ROIs = caiman_datadir() + '/example_movies/volpy/gui_roi.hdf5'
with h5py.File(path_ROIs, 'r') as fl:
ROIs = fl['mov'][()]
elif method == 'maskrcnn': # Important!! make sure install keras before using mask rcnn
weights_path = download_model('mask_rcnn')
weights_path = '/home/nel/Code/NEL_LAB/Mask_RCNN/logs/neurons20200824T1032/mask_rcnn_neurons_0040.h5'
ROIs = utils.mrcnn_inference(
img=summary_images.transpose([1, 2, 0]),
size_range=[5, 100],
weights_path=weights_path,
display_result=True
) # size parameter decides size range of masks to be selected
#np.save(os.path.join(file_dir, 'ROIs'), ROIs)
# %% restart cluster to clean up memory
cm.stop_server(dview=dview)
c, dview, n_processes = cm.cluster.setup_cluster(backend='local',
n_processes=None,
single_thread=False,
maxtasksperchild=1)
# %% parameters for trace denoising and spike extraction
ROIs = ROIs # region of interests
index = list(range(len(ROIs))) # index of neurons
weights = None # reuse spatial weights
context_size = 35 # number of pixels surrounding the ROI to censor from the background PCA
flip_signal = True # Important!! Flip signal or not, True for Voltron indicator, False for others
hp_freq_pb = 1 / 3 # parameter for high-pass filter to remove photobleaching
threshold_method = 'adaptive_threshold' # 'simple' or 'adaptive_threshold'
min_spikes = 30 # minimal spikes to be found
threshold = 4 # threshold for finding spikes, increase threshold to find less spikes
do_plot = False # plot detail of spikes, template for the last iteration
ridge_bg = 0.01 # ridge regression regularizer strength for background removement, larger value specifies stronger regularization
sub_freq = 20 # frequency for subthreshold extraction
weight_update = 'ridge' # 'ridge' or 'NMF' for weight update
n_iter = 2
opts_dict = {
'fnames': fname_new,
'ROIs': ROIs,
'index': index,
'weights': weights,
'context_size': context_size,
'flip_signal': flip_signal,
'hp_freq_pb': hp_freq_pb,
'threshold_method': threshold_method,
'min_spikes': min_spikes,
'threshold': threshold,
'do_plot': do_plot,
'ridge_bg': ridge_bg,
'sub_freq': sub_freq,
'weight_update': weight_update,
'n_iter': n_iter
}
opts.change_params(params_dict=opts_dict)
if options is not None:
print('using external options')
opts.change_params(params_dict=options)
else:
print('not using external options')
#%% TRACE DENOISING AND SPIKE DETECTION
vpy = VOLPY(n_processes=n_processes, dview=dview, params=opts)
vpy.fit(n_processes=n_processes, dview=dview)
#%% visualization
display_images = False
if display_images:
print(np.where(
vpy.estimates['locality'])[0]) # neurons that pass locality test
idx = np.where(vpy.estimates['locality'] > 0)[0]
utils.view_components(vpy.estimates, img_corr, idx)
#%% reconstructed movie
# note the negative spatial weights is cutoff
if display_images:
mv_all = utils.reconstructed_movie(vpy.estimates,
fnames=mc.mmap_file,
idx=idx,
scope=(0, 1000),
flip_signal=flip_signal)
mv_all.play(fr=40)
#%% save the result in .npy format
save_result = True
if save_result:
vpy.estimates['ROIs'] = ROIs
save_name = f'volpy_{os.path.split(fnames)[1][:-5]}_{opts.volspike["threshold_method"]}_{opts.volspike["threshold"]}_{opts.volspike["weight_update"]}_bg_{opts.volspike["ridge_bg"]}'
np.save(os.path.join(file_dir, save_name), vpy.estimates)
# %% STOP CLUSTER and clean up log files
cm.stop_server(dview=dview)
log_files = glob.glob('*_LOG_*')
for log_file in log_files:
os.remove(log_file)
def main():
pass # For compatibility between running under Spyder and the CLI
# %% Load demo movie and ROIs
fnames = download_demo(
'demo_voltage_imaging.hdf5',
'volpy') # file path to movie file (will download if not present)
path_ROIs = download_demo(
'demo_voltage_imaging_ROIs.hdf5',
'volpy') # file path to ROIs file (will download if not present)
file_dir = os.path.split(fnames)[0]
#%% dataset dependent parameters
# dataset dependent parameters
fr = 400 # sample rate of the movie
# motion correction parameters
pw_rigid = False # flag for pw-rigid motion correction
gSig_filt = (3, 3) # size of filter, in general gSig (see below),
# change this one if algorithm does not work
max_shifts = (5, 5) # maximum allowed rigid shift
strides = (
48, 48
) # start a new patch for pw-rigid motion correction every x pixels
overlaps = (24, 24
) # overlap between pathes (size of patch strides+overlaps)
max_deviation_rigid = 3 # maximum deviation allowed for patch with respect to rigid shifts
border_nan = 'copy'
opts_dict = {
'fnames': fnames,
'fr': fr,
'pw_rigid': pw_rigid,
'max_shifts': max_shifts,
'gSig_filt': gSig_filt,
'strides': strides,
'overlaps': overlaps,
'max_deviation_rigid': max_deviation_rigid,
'border_nan': border_nan
}
opts = volparams(params_dict=opts_dict)
# %% play the movie (optional)
# playing the movie using opencv. It requires loading the movie in memory.
# To close the movie press q
display_images = False
if display_images:
m_orig = cm.load(fnames)
ds_ratio = 0.2
moviehandle = m_orig.resize(1, 1, ds_ratio)
moviehandle.play(q_max=99.5, fr=40, magnification=4)
# %% start a cluster for parallel processing
c, dview, n_processes = cm.cluster.setup_cluster(backend='local',
n_processes=None,
single_thread=False)
# %%% MOTION CORRECTION
# first we create a motion correction object with the specified parameters
mc = MotionCorrect(fnames, dview=dview, **opts.get_group('motion'))
# Run correction
do_motion_correction = True
if do_motion_correction:
mc.motion_correct(save_movie=True)
else:
mc_list = [
file for file in os.listdir(file_dir)
if (os.path.splitext(os.path.split(fnames)[-1])[0] in file
and '.mmap' in file)
]
mc.mmap_file = [os.path.join(file_dir, mc_list[0])]
print(f'reuse previously saved motion corrected file:{mc.mmap_file}')
# %% compare with original movie
if display_images:
m_orig = cm.load(fnames)
m_rig = cm.load(mc.mmap_file)
ds_ratio = 0.2
moviehandle = cm.concatenate(
[m_orig.resize(1, 1, ds_ratio),
m_rig.resize(1, 1, ds_ratio)],
axis=2)
moviehandle.play(fr=40, q_max=99.5, magnification=4) # press q to exit
# %% MEMORY MAPPING
do_memory_mapping = True
if do_memory_mapping:
border_to_0 = 0 if mc.border_nan == 'copy' else mc.border_to_0
# you can include the boundaries of the FOV if you used the 'copy' option
# during motion correction, although be careful about the components near
# the boundaries
# memory map the file in order 'C'
fname_new = cm.save_memmap_join(
mc.mmap_file,
base_name='memmap_' +
os.path.splitext(os.path.split(fnames)[-1])[0],
add_to_mov=border_to_0,
dview=dview) # exclude border
else:
mmap_list = [
file for file in os.listdir(file_dir)
if ('memmap_' +
os.path.splitext(os.path.split(fnames)[-1])[0]) in file
]
fname_new = os.path.join(file_dir, mmap_list[0])
print(f'reuse previously saved memory mapping file:{fname_new}')
# %% SEGMENTATION
# create summary images
img = mean_image(mc.mmap_file[0], window=1000, dview=dview)
img = (img - np.mean(img)) / np.std(img)
gaussian_blur = False # Use gaussian blur when there is too much noise in the video
Cn = local_correlations_movie_offline(mc.mmap_file[0],
fr=fr,
window=fr * 4,
stride=fr * 4,
winSize_baseline=fr,
remove_baseline=True,
gaussian_blur=gaussian_blur,
dview=dview).max(axis=0)
img_corr = (Cn - np.mean(Cn)) / np.std(Cn)
summary_images = np.stack([img, img, img_corr], axis=0).astype(np.float32)
# save summary images which are used in the VolPy GUI
cm.movie(summary_images).save(fnames[:-5] + '_summary_images.tif')
fig, axs = plt.subplots(1, 2)
axs[0].imshow(summary_images[0])
axs[1].imshow(summary_images[2])
axs[0].set_title('mean image')
axs[1].set_title('corr image')
#%% methods for segmentation
methods_list = [
'manual_annotation', # manual annotations need prepared annotated datasets in the same format as demo_voltage_imaging_ROIs.hdf5
'maskrcnn', # Mask R-CNN is a convolutional neural network trained for detecting neurons in summary images
'gui_annotation'
] # use VolPy GUI to correct outputs of Mask R-CNN or annotate new datasets
method = methods_list[0]
if method == 'manual_annotation':
with h5py.File(path_ROIs, 'r') as fl:
ROIs = fl['mov'][()]
elif method == 'maskrcnn': # Important!! Make sure install keras before using mask rcnn.
weights_path = download_model(
'mask_rcnn'
) # also make sure you have downloaded the new weight. The weight was updated on Dec 1st 2020.
ROIs = utils.mrcnn_inference(
img=summary_images.transpose([1, 2, 0]),
size_range=[5, 22],
weights_path=weights_path,
display_result=True
) # size parameter decides size range of masks to be selected
cm.movie(ROIs).save(fnames[:-5] + 'mrcnn_ROIs.hdf5')
elif method == 'gui_annotation':
# run volpy_gui.py file in the caiman/source_extraction/volpy folder
gui_ROIs = caiman_datadir() + '/example_movies/volpy/gui_roi.hdf5'
with h5py.File(gui_ROIs, 'r') as fl:
ROIs = fl['mov'][()]
fig, axs = plt.subplots(1, 2)
axs[0].imshow(summary_images[0])
axs[1].imshow(ROIs.sum(0))
axs[0].set_title('mean image')
axs[1].set_title('masks')
# %% restart cluster to clean up memory
cm.stop_server(dview=dview)
c, dview, n_processes = cm.cluster.setup_cluster(backend='local',
n_processes=None,
single_thread=False,
maxtasksperchild=1)
# %% parameters for trace denoising and spike extraction
ROIs = ROIs # region of interests
index = list(range(len(ROIs))) # index of neurons
weights = None # reuse spatial weights
context_size = 35 # number of pixels surrounding the ROI to censor from the background PCA
visualize_ROI = False # whether to visualize the region of interest inside the context region
flip_signal = True # Important!! Flip signal or not, True for Voltron indicator, False for others
hp_freq_pb = 1 / 3 # parameter for high-pass filter to remove photobleaching
clip = 100 # maximum number of spikes to form spike template
threshold_method = 'adaptive_threshold' # adaptive_threshold or simple
min_spikes = 10 # minimal spikes to be found
pnorm = 0.5 # a variable deciding the amount of spikes chosen for adaptive threshold method
threshold = 3 # threshold for finding spikes only used in simple threshold method, Increase the threshold to find less spikes
do_plot = False # plot detail of spikes, template for the last iteration
ridge_bg = 0.01 # ridge regression regularizer strength for background removement, larger value specifies stronger regularization
sub_freq = 20 # frequency for subthreshold extraction
weight_update = 'ridge' # ridge or NMF for weight update
n_iter = 2 # number of iterations alternating between estimating spike times and spatial filters
opts_dict = {
'fnames': fname_new,
'ROIs': ROIs,
'index': index,
'weights': weights,
'context_size': context_size,
'visualize_ROI': visualize_ROI,
'flip_signal': flip_signal,
'hp_freq_pb': hp_freq_pb,
'clip': clip,
'threshold_method': threshold_method,
'min_spikes': min_spikes,
'pnorm': pnorm,
'threshold': threshold,
'do_plot': do_plot,
'ridge_bg': ridge_bg,
'sub_freq': sub_freq,
'weight_update': weight_update,
'n_iter': n_iter
}
opts.change_params(params_dict=opts_dict)
#%% TRACE DENOISING AND SPIKE DETECTION
vpy = VOLPY(n_processes=n_processes, dview=dview, params=opts)
vpy.fit(n_processes=n_processes, dview=dview)
#%% visualization
display_images = True
if display_images:
print(np.where(
vpy.estimates['locality'])[0]) # neurons that pass locality test
idx = np.where(vpy.estimates['locality'] > 0)[0]
utils.view_components(vpy.estimates, img_corr, idx)
#%% reconstructed movie
# note the negative spatial weights is cutoff
if display_images:
mv_all = utils.reconstructed_movie(vpy.estimates.copy(),
fnames=mc.mmap_file,
idx=idx,
scope=(0, 1000),
flip_signal=flip_signal)
mv_all.play(fr=40)
#%% save the result in .npy format
save_result = True
if save_result:
vpy.estimates['ROIs'] = ROIs
vpy.estimates['params'] = opts
save_name = f'volpy_{os.path.split(fnames)[1][:-5]}_{threshold_method}'
np.save(os.path.join(file_dir, save_name), vpy.estimates)
# %% STOP CLUSTER and clean up log files
cm.stop_server(dview=dview)
log_files = glob.glob('*_LOG_*')
for log_file in log_files:
os.remove(log_file)
