Bilateral Pathways from the Basal Forebrain to Sensory Cortices May Contribute to Synchronous Sensory Processing

Abstract

Sensory processing in the cortex should integrate inputs arriving from receptive fields located on both sides of the body. This role could be played by the corpus callosum through precise projections between both hemispheres. However, different studies suggest that cholinergic projections from the basal forebrain (BF) could also contribute to the synchronization and integration of cortical activities. Using tracer injections and optogenetic techniques in transgenic mice, we investigated whether the BF cells project bilaterally to sensory cortical areas, and have provided anatomical evidence to support a modulatory role for the cholinergic projections in sensory integration. Application of the retrograde tracer Fluor-Gold or Fast Blue in both hemispheres of the primary somatosensory (S1), auditory or visual cortical areas showed labeled neurons in the ipsi- and contralateral areas of the diagonal band of Broca and substantia innominata. The nucleus basalis magnocellularis only showed ipsilateral projections to the cortex. Optogenetic stimulation of the horizontal limb of the diagonal band of Broca facilitated whisker responses in the S1 cortex of both hemispheres through activation of muscarinic cholinergic receptors and this effect was diminished by atropine injection. In conclusion, our findings have revealed that specific areas of the BF project bilaterally to sensory cortices and may contribute to the coordination of neuronal activity on both hemispheres.

Keywords: basal magnocellular nucleus; cholinergic neurons; diagonal band of Broca; mouse; optogenetic stimulation; somatosensory evoked potential.

Figure 1

Location of injection sites into primary somatosensory (S1) and primary auditory (A1) cortices. (A) Schematic drawing and microphotograph of a brain coronal section showing the injection site in A1 cortex. (B) The injection site is located in S1 cortex.

Figure 2

Confocal microphotograph of vertical limb of the diagonal band of Broca (HDB) showing labeled neurons of animals injected with the fluorescent tracers in the left hemisphere in S1 (yellow neurons) and A1 (blue neurons). Since the injection was in the left hemisphere, ipsilateral neurons are at the left side of the dash line (indicated above) and contralateral neurons are the ones located at the right side of the dash line indicating the middle line (upper images). Bottom left image shows positive neurons for choline-acetyltransferase (ChAT) immunostaining labeled in red color. FB: Fast Blue (blue) labeled neurons; Fluoro-Gold (FlGo; yellow): FlGo labeled neurons; ChAT (red): cholinergic labeled neurons. Labelled neurons were observed both in the ipsi (left hemisphere) and the contralateral (right) injection site.

Figure 3

Confocal microphotographs of labeled neurons in horizontal limb of the diagonal band of Broca (HDB) nuclei of the basal forebrain (BF) of animal injected with both fluorescent tracers in the left hemisphere in S1 (yellow neurons) and A1 (blue neurons). The left column shows the HDB labeled neurons of the ipsilateral injection site; the right column shows the HDB contralateral labeled neurons to the injection site. Blue indicated FB labeled neurons; Yellow indicated FlGo labeled neurons and red color indicates positive neurons for ChAT immunostaining. The upper row shows the merge of all labeled neurons in HDB.

Figure 4

Fluorescent microscope images showing the rostro-caudal distribution of fluorescent labeled neurons. (A–C) Fluorescent microscope images from animals injected with FlGo and FB in S1. (D–F) Fluorescent microscope images from animals injected with FlGo in A1 and FB deposit in A1. (G–I) Fluorescent microscope images from animals injected with FlGo in V1 and FB deposit in V1. (J–L) Graphic representation of rostro-caudal distribution of neurons in VDB, HDB and B nucleus and the proportion of contralateral labeled neurons.

Figure 5

Effect of blue-light stimulation of HDB and B nuclei on ipsi- and contralateral S1 cortices. (A) Plot of the somatosensory evoked potential (SEP) area during the control period (4 min before blue-light stimulation) and 30 min after blue light stimulation of HDB. The mean area of the control period was considered as 100%. HDB induced a facilitation of both SEP recorded in both hemispheres, although the area increased slower in the contralateral cortex to the stimulated HDB. (B) Same plot as in (A) after blue light stimulation of B nucleus. Note that contralateral SEP area was not affected. (C) Plots of the effect of atropine sulfate on HDB stimulation. The effect was measured 10 min after HDB stimulation respect to the mean SEP area during the control period (4 min before blue light stimulation). After saline i.p. injection blue-light stimulation induced a facilitation of SEP area in both ipsi- and contralateral cortices (n = 8). However, the effect was blocked when atropine was i.p. injected 10 min before HDB stimulation. Insets in (A,B) show representative traces of the SEP in control and 10 min after optogenetic stimulation (black and blue tracers, respectively). *p < 0.05; **p < 0.01.