Optophysiological analysis of associational circuits in the olfactory cortex

Primary olfactory cortical areas receive direct input from the olfactory bulb, but also have extensive associational connections that have been mainly studied with classical anatomical methods. Here, we shed light on the functional properties of associational connections in the anterior and posterior piriform cortices (aPC and pPC) using optophysiological methods. We found that the aPC receives dense functional connections from the anterior olfactory nucleus (AON), a major hub in olfactory cortical circuits. The local recurrent connectivity within the aPC, long invoked in cortical autoassociative models, is sparse and weak. By contrast, the pPC receives negligible input from the AON, but has dense connections from the aPC as well as more local recurrent connections than the aPC. Finally, there are negligible functional connections from the pPC to aPC. Our study provides a circuit basis for a more sensory role for the aPC in odor processing and an associative role for the pPC

LED Arrays as Cost Effective and Efficient Light Sources for Widefield Microscopy

New developments in fluorophores as well as in detection methods have fueled the rapid growth of optical imaging in the life sciences. Commercial widefield microscopes generally use arc lamps, excitation/emission filters and shutters for fluorescence imaging. These components can be expensive, difficult to maintain and preclude stable illumination. Here, we describe methods to construct inexpensive and easy-to-use light sources for optical microscopy using light-emitting diodes (LEDs). We also provide examples of its applicability to biological fluorescence imaging

Optophysiological analysis of associational circuits in the olfactory cortex

Primary olfactory cortical areas receive direct input from the olfactory bulb, but also have extensive associational connections that have been mainly studied with classical anatomical methods. Here, we shed light on the functional properties of associational connections in the anterior and posterior piriform cortex (aPC and pPC) using optophysiological methods. We found that the aPC receives dense functional connections from the anterior olfactory nucleus (AON), a major hub in olfactory cortical circuits. The local recurrent connectivity within the aPC, long invoked in cortical autoassociative models, is sparse and weak. By contrast, the pPC receives negligible input from the AON, but has dense connections from the aPC as well as more local recurrent connections than the aPC. Finally, there are negligible functional connections from the pPC to aPC. Our study provides a circuit basis for a more sensory role for the aPC in odor processing and an associative role for the pPC

Pulsed ultrasound differentially stimulates somatosensory circuits in humans as indicated by EEG and FMRI.

Peripheral somatosensory circuits are known to respond to diverse stimulus modalities. The energy modalities capable of eliciting somatosensory responses traditionally belong to mechanical, thermal, electromagnetic, and photonic domains. Ultrasound (US) applied to the periphery has also been reported to evoke diverse somatosensations. These observations however have been based primarily on subjective reports and lack neurophysiological descriptions. To investigate the effects of peripherally applied US on human somatosensory brain circuit activity we recorded evoked potentials using electroencephalography and conducted functional magnetic resonance imaging of blood oxygen level-dependent (BOLD) responses to fingertip stimulation with pulsed US. We found a pulsed US waveform designed to elicit a mild vibration sensation reliably triggered evoked potentials having distinct waveform morphologies including a large double-peaked vertex potential. Fingertip stimulation with this pulsed US waveform also led to the appearance of BOLD signals in brain regions responsible for somatosensory discrimination including the primary somatosensory cortex and parietal operculum, as well as brain regions involved in hierarchical somatosensory processing, such as the insula, anterior middle cingulate cortex, and supramarginal gyrus. By changing the energy profile of the pulsed US stimulus waveform we observed pulsed US can differentially activate somatosensory circuits and alter subjective reports that are concomitant with changes in evoked potential morphology and BOLD response patterns. Based on these observations we conclude pulsed US can functionally stimulate different somatosensory fibers and receptors, which may permit new approaches to the study and diagnosis of peripheral nerve injury, dysfunction, and disease

A standalone two wavelength illuminator.

<p>Two different LEDs, with independent electronic driving circuitry similar to that shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0002146#pone-0002146-g001" target="_blank">Figure 1</a> are used. A dichroic mirror passes light from one LED and reflects light from the other onto the end of the fiber light guide. Any necessary excitation filters can be placed on the LEDs.</p

Two methods for coupling an LED based illuminator to a microscope.

<p>(A) An optical is fiber placed on the surface of an LED to direct light into a small box consisting of a focusing lens and mirror that is inserted into a side port on the microscope. (B) A homemade “lamp” consisting of an LED, heatsink, tube, and lens is coupled to the lamp housing port in the back of the microscope.</p

A high intensity Luxeon V LED and driving circuitry.

<p>(A) A high intensity 5 W Luxeon V blue LED is shown mounted on a standard fin type heatsink with thermal grease and an electrical insulator in between. There are no moving parts used, reducing susceptibility to mechanical damage. (B) A simple electronic circuit that can be used to drive a single blue LED is shown. Its output intensity is adjusted by a potentiometer; the light can be turned on and off by a low current TTL input. (C) An LED driven by a high square wave input follows the signal closely. The speed of a typical LED array is clearly illustrated by its response to a voltage step. The driver circuit was that shown in B.</p

A custom built widefield microscope.

<p>(A) As an alternative to a standard commercial microscope, we used a custom built setup as shown above for our <i>in vivo</i> widefield experiments. A CCD camera is coupled to two lenses along with optional extension tubes to adjust the magnification. The ordering of parts is as follows: CCD camera, an F to C mount adapter with an emission filter placed inside or on top of the coupler, a 62 mm, 105 mm FL lens, a 62 mm to 52 mm step down ring, a 52 mm to 52 mm coupling ring, and a 52 mm, 50 mm FL lens. (B) Shown here are sample individual frames of the dorsal surface of the olfactory bulb, under white light, revealing the blood vessel architecture, as well as resting synaptopHluorin and intrinsic optical signals from the same animal respectively under blue and far red illumination. (C) The changes in spH and intrinsic signals upon presentation of fresh air or two odorants. Images show fractional changes (ΔF/F) with increases shown by brighter pixels and decreases by darker pixels.</p

Comparison of an LED based illuminator and Xenon arc lamp.

<p>(A) Images of hippocampal astrocytes in cell culture labeled with antibodies against the astrocytic marker GFAP are shown for illumination with Xenon lamp and with LED. (B) Time traces of fluorescence intensity of the same region of the field (shown by white square) in the two conditions. (C) Distribution of pixel intensities of the images.</p