1. Volume 138, Issue 1, Pages 266-274 (January 2010)
The β3-Adrenoceptor Agonist GW (Solabegron) Decreases Excitability of Human Enteric Neurons via Release of Somatostatin 
Michael Schemann, Nadia Hafsi, Klaus Michel, Olivia I. Kober, Jutta Wollmann, Qin Li, Florian Zeller, Rupert Langer, Kevin Lee, Selim Cellek 
Gastroenterology 
Volume 138, Issue 1, Pages (January 2010)
DOI: /j.gastro
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2. Figure 1
GW had no immediate postsynaptic effects in human submucous neurons but decreased fast synaptic input. (A) An unfiltered trace recorded from a neuron with the CCD camera shows slow changes in membrane potential. The steady increase in baseline is due to bleaching of the voltage-sensitive dye and is not related to changes in membrane potential. There is no change in the baseline during or after spritz application of GW (indicated by the bar), suggesting that GW did not cause immediate hyperpolarization or depolarization of the membrane potential. Interganglionic fiber tracts were electrically stimulated shortly before and after GW application (arrows) evoked compound action potentials (first spike) and synaptically mediated fast EPSP-triggered action potentials (late spike). Expanded signals from a different neuron are seen (B). Although the compound action potential (first spike) remained unchanged, GW reduced the fast EPSP-triggered action potentials (late spikes; left trace before right trace after GW application). (C) A β3-AR labeling in a submucous ganglion clearly shows staining in neuronal somata (one marked by arrowhead) as well as labeling of nerve varicosities (2 marked by arrows). The staining in the soma appears to be cytoplasmic (see Supplementary Video).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2010 AGA Institute Terms and Conditions

3. Figure 2
GW decreased excitability in human submucous neurons. (A) Traces show nicotine-evoked spike discharge in a submucous neuron. Bars below the traces mark nicotine application. In the presence of 0.5 μM GW nicotine-evoked spike discharge is decreased. After 1-hour washout the nicotine response recovered. (B) Summary of control responses to nicotine which are significantly reduced in 0.1 μM, 0.3 μM, 0.5 μM, and 1 μM GW Asterisks indicate significant changes. Also shown is partial recovery after washout.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2010 AGA Institute Terms and Conditions

4. Figure 3
Inhibitory actions of GW were reversed by β3 AR antagonist SR (A) Consecutive traces from 1 neuron top to bottom are shown. Nicotine application is indicated by the bars below the traces. Initially, a nicotine spritz application evoked a moderate spike discharge which is enhanced in the presence of the β-3 receptor antagonist SR The addition of 0.5 μM GW did not produce any inhibition of the nicotine response. After washout there is still an enhanced response to nicotine which is reduced by perfusion of 0.5 μM GW (B) Summary of such experiments in 43 neurons. SR significantly enhanced nicotine-evoked spike discharge. In the presence of SR-59230, GW did not inhibit the nicotine-evoked spike discharge. After washout of >1 hour the nicotine response is still enhanced, but now the addition of GW significantly reduced nicotine-evoked spike discharge.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2010 AGA Institute Terms and Conditions

5. Figure 4
Inhibitory effect of GW on nicotine-evoked spike discharge is prevented by SST2 receptor antagonism. (A) A neuron that responded to nicotine spritz application (marked by the bars) is shown. The spike discharge under control conditions is almost completely blocked after 20-minute perfusion of 0.5 μM GW The addition of the SST2 receptor antagonist CYN partially reversed this inhibitory effect. (B) Summary of the results from 18 neurons. GW significantly reduced spike frequency. The addition of CYN partially reversed this inhibition with a significant increase in nicotine-evoked spike discharge.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2010 AGA Institute Terms and Conditions

6. Figure 5
Immunohistochemistry in whole mounts of the human submucous plexus layer. Adipocytes in the human submucous plexus layer show adiponectin immunoreactivity (A). Immunoreactivity is seen as a thin peripheral ring because of the dominating intracellular fat droplet. Double labeling showed that the 3 adipocytes marked by arrowheads are also β3-AR positive (B). The 2 arrows mark interganglionic fiber tracts of the submucous plexus. SST immunoreactivity in adipocytes is shown by the ring-like staining (C, some are marked by arrowheads). The arrow is pointing at an interganglionic fiber tract that is out of focus. (D) Double labeling is shown of adiponectin-positive adipocytes (green) and TH-positive nerve fibers (red). Note the varicose TH nerve in close proximity to the adipocytes. (Bottom right) A blood vessel innervated by TH-positive fibers is shown. Although the TH-positive fibers are likely of sympathetic origin, we regularly observed TH-positive nerve cell bodies in submucous ganglia (E). (F) A submucous ganglion labeled with the pan neuronal marker PGP 9.5 is shown. Double labeling of the same ganglion showed dense SST2-receptor staining in the ganglion. Two neurons that are SST2-receptor negative are labeled by arrowheads.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2010 AGA Institute Terms and Conditions

7. Figure 6
GW increased SST release from human primary adipocytes. The effect of β3-AR agonist GW (1 nM to 10 μM) on SST and adiponectin release from human primary adipocytes in the absence (control) or presence of 1 μM SR59230 (β3-AR antagonist); each column represents the mean of 3 experiments; error bars represent the standard error of mean.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2010 AGA Institute Terms and Conditions