#SFN18 Day 2 Recap: Controlling Neurons with Ultrasound and a Novel Avenue for Depression Treatment?

Sonogenetics - A non-invasive method to manipulate neurons

During the AM poster sessions, one that caught my eye was from the Chalasani Lab at the Salk Institute in La Jolla, California. Several years ago, they described a method by which they could control neural activity in the nematode worm C. elegans using focused ultrasound. This paper demonstrated that ectopic expression of the mechanosensitive channel TRP-4 in neurons rendered them sensitive to ultrasound stimulation. This is a big deal because other so called ‘non-invasive’ neural manipulation techniques like optogenetics require a fiber optic probe to be placed near the cells of interest, making the manipulation of deep brain structures with high temporal precision tedious.

Sonogenetics allows for non-invasive control of neural activity. Here, in C. elegans with PVD neurons expressing the ultrasound-sensitive protein (TRP-4) and the calcium indicator GCaMP3, we can see that ultrasound exposure drastically increases calcium activity in these neurons, indicating ultrasound mediated neural activation. Warmer colors indicate more GCaMP3 fluorescence = more activity  (Credit: Ibsen et al., 2015;  Nature Communications )

Sonogenetics allows for non-invasive control of neural activity. Here, in C. elegans with PVD neurons expressing the ultrasound-sensitive protein (TRP-4) and the calcium indicator GCaMP3, we can see that ultrasound exposure drastically increases calcium activity in these neurons, indicating ultrasound mediated neural activation. Warmer colors indicate more GCaMP3 fluorescence = more activity (Credit: Ibsen et al., 2015; Nature Communications)

This was to be just the first step in a long process of isolating different mechanosensitive proteins and screening them in mammalian cells to find one just right for use in more complicated organisms. During the poster session this morning, Corinne Lee-Kubli, a post-doc in the Chalasani lab, provided an update on the progress in sonogenetics to date.

Using an in vitro screening method to identify ultrasound-sensitive mechanoreceptors, Corinne expressed a large variety of putative channels in cells in a dish. These cells were co-transfected with the calcium indicator GCaMP6f, a powerful and fast reporter of cell activity. The fluorescent signal was then monitored before, during, and after ultrasound stimulation in a high-throughput manner.

A subset of the putative mechanoreceptors were packaged into cre-dependent AAV-viral vectors and delivered to AgRP neurons deep in the brain (arcuate nucleus) of AgRP-cre mice. Validation of the excitatory actions of the ultrasound sensitive protein was done using a feeding assay, as AgRP neurons strongly promote feeding. Upon ultrasound (10 MHz) stimulation of the head (through the skull and entire brain), a few of the channels strongly promoted feeding responses, a trait not observed in mice expressing the control virus (encoding GFP). An important note is that ultrasound stimulation alone had no effect on feeding responses, indicating a specific effect of the putative mechanoreceptor in AgRP neurons.

AgRP neurons in the arcuate nucleus expressing the calcium indicator GCaMP6. These cells are powerful regulators of feeding behavior and metabolism  ( Credit: Srisai et al., 2017 ;  Nature Communications )

AgRP neurons in the arcuate nucleus expressing the calcium indicator GCaMP6. These cells are powerful regulators of feeding behavior and metabolism (Credit: Srisai et al., 2017; Nature Communications)

This proof-of-principle application represents a significant advancement for the nascent field of sonogenetics. Much more research needs to be done to discover the most potent and specific ultrasound sensitive protein, the kinetics of said protein, and additional tools for cell inhibition. In the future, we can expect to see multiple channels expressed in different cellular populations, each sensitive to different ultrasound frequencies. Then, ‘nested’ delivery of different ultrasound waveforms could putatively activate and/or inhibit discrete cell populations across the entire brain, simultaneously or with tight temporal control.

The power of this technique is impressive, as ultrasound can easily reach through the entire mouse brain at 10 MHz, and can go much deeper (e.g., in rat or primate brain) using lower frequencies. I look forward to what’s to come!

Bidirectional Control of Depression Through Hypothalamic Feeding Circuits

Speaking of the arcuate nucleus, during the day 2 poster sessions, one that caught my eye was titled “Chronic unpredictable stress modulates neuronal activity of AgRP and POMC neurons in hypothalamic arcuate nucleus” presented by Xing Fang in the Xin-yun Lu lab at the Medical College of Georgia at Augusta University.

Agouti-related peptide (AgRP) and pro-opiomelanocortin (POMC) neurons in the arcuate nucleus strongly regulate feeding behavior and food intake. Broadly, AgRP neurons promote feeding (orexigenic), while POMC neurons work in a reciprocal manner to suppress feeding (anorexigenic).

AgRP neurons in the arcuate promote food intake while POMC neurons inhibit food intake via their actions on downstream MC4R- expressing neurons in the paraventricular nucleus  (Credit: Carol A. Rouzer, Vanderbilt University)

AgRP neurons in the arcuate promote food intake while POMC neurons inhibit food intake via their actions on downstream MC4R- expressing neurons in the paraventricular nucleus (Credit: Carol A. Rouzer, Vanderbilt University)

Depression is characterized by aberrant responses to environmental stimuli. For example, chronic psychological stress can promote depression in humans and animal models. Stress-induced depression is characterized by anhedonia (not enjoying what you used to love), lethargy and despair, and changes in feeding behavior and appetite. How does stress cause these behaviors to come about?

Using in vivo electrophysiology, behavioral assays, and DREADDs, Fang and colleagues investigated the role of hypothalamic AgRP and POMC neurons (two populations that powerfully control appetite) in mediating these behaviors.

This work builds on previous studies by the group, long linking depressive-like behavior to alterations in feeding and satiety hormones such as leptin.

To induce depression in mice, the researchers used a technique called ‘chronic unpredictable stress’ (CUS). This model strongly promotes a depression-like state after 10 days of unpredictable stress where mice go through a gamut of constant light exposure, tail pinches, restraint, and shock stimuli, among others.

Viral injections into the arcuate nucleus of POMC-Cre mice (left panels; projections in red) shows their wide axonal distribution throughout the brain. Similarly, injections into the arcuate nucleus of AgRP-cre mice demonstrate that they also project throughout the brain, although in a different pattern (right panels, projections in green)  (Credit:  Wang et al., 2015 ;  Frontiers in Neuroanatomy )

Viral injections into the arcuate nucleus of POMC-Cre mice (left panels; projections in red) shows their wide axonal distribution throughout the brain. Similarly, injections into the arcuate nucleus of AgRP-cre mice demonstrate that they also project throughout the brain, although in a different pattern (right panels, projections in green) (Credit: Wang et al., 2015; Frontiers in Neuroanatomy)

Through their electrophysiological recordings, the researchers demonstrated that CUS decreased the firing rate of AgRP neurons but increased the firing rate of POMC neurons. When they tested the role of AgRP neurons in depressive-like behavior using stimulatory (Gq) or inhibitory (Gi) DREADDs, they were able to elicit opposite responses. Stimulation of these neurons improved depressive-like behavior, while inhibition promoted it.

Together, these studies suggest that AgRP and POMC neurons play an important role in stress-related adaptive behavior. Importantly, they provide a novel circuit related to depression which may be targetable for the treatment of the disease through pharmacological agents or lifestyle changes.

That’s all from me for day two, where I tried to focus more on posters than talks! Unfortunately I could only highlight 2 out of >5,000 interesting ones!

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