Dittrich K et al. (2014), Purinergic receptor-induced Ca2+ signaling in t...

XB-ART-47656

Purinergic Signal 2014 Jan 01;102:327-36. doi: 10.1007/s11302-013-9402-3.

Show Gene links Show Anatomy links

Purinergic receptor-induced Ca2+ signaling in the neuroepithelium of the vomeronasal organ of larval Xenopus laevis.

Dittrich K , Sansone A , Hassenklöver T , Manzini I .

???displayArticle.abstract???
Purinergic signaling has considerable impact on the functioning of the nervous system, including the special senses. Purinergic receptors are expressed in various cell types in the retina, cochlea, taste buds, and the olfactory epithelium. The activation of these receptors by nucleotides, particularly adenosine-5'-triphosphate (ATP) and its breakdown products, has been shown to tune sensory information coding to control the homeostasis and to regulate the cell turnover in these organs. While the purinergic system of the retina, cochlea, and taste buds has been investigated in numerous studies, the available information about purinergic signaling in the olfactory system is rather limited. Using functional calcium imaging, we identified and characterized the purinergic receptors expressed in the vomeronasal organ of larval Xenopus laevis. ATP-evoked activity in supporting and basal cells was not dependent on extracellular Ca(2+). Depletion of intracellular Ca(2+) stores disrupted the responses in both cell types. In addition to ATP, supporting cells responded also to uridine-5'-triphosphate (UTP) and adenosine-5'-O-(3-thiotriphosphate) (ATPγS). The response profile of basal cells was considerably broader. In addition to ATP, they were activated by ADP, 2-MeSATP, 2-MeSADP, ATPγS, UTP, and UDP. Together, our findings suggest that supporting cells express P2Y(2)/P2Y(4)-like purinergic receptors and that basal cells express multiple P2Y receptors. In contrast, vomeronasal receptor neurons were not sensitive to nucleotides, suggesting that they do not express purinergic receptors. Our data provide the basis for further investigations of the physiological role of purinergic signaling in the vomeronasal organ and the olfactory system in general.

???displayArticle.pubmedLink??? 24271060
???displayArticle.pmcLink??? PMC4040167
???displayArticle.link??? Purinergic Signal

Species referenced: Xenopus laevis
Genes referenced: abcc8 cpa1 krt12.4 tal1 znf469

???attribute.lit??? ???displayArticles.show???

	Fig. 1. Morphological organization of the epithelium of the vomeronasal organ of larval Xenopus laevis. a Slice of a VNO with biocytin-streptavidin-stained VRNs (SCL supporting cell layer; VRNL vomeronasal receptor neuron layer; BCL basal cell layer). b Supporting cell staining with an antibody against cytokeratin type II of the same slice. c Overlay of VRNs (a red fluorescence) and SCs (b green fluorescence). Basal cells are biocytin-streptavidin and cytokeratin II negative and are located in the BCL. d and e Higher magnification of the region delineated by a white rectangle in a and b, respectively. The filled arrowheads in d indicate the somata of some VRNs. The open arrowheads indicate the apical endings of some VRNs (knobs). The open arrowheads in e indicate the soma of a SC, the filled arrowheads point at the basal process of the SC spanning the whole epithelium of the VNO. f Slice of another VNO with propidium iodide-stained cell nuclei (red fluorescence) and BrdU-positive cells (green fluorescence). The vast majority of BrdU positive cells are confined to the BCL of the VNO. g Higher magnification of the region delineated by a white rectangle in f. The filled arrowheads indicate BrdU-positive cells in the basalmost part of the epithelium. Asterisks indicate BrdU-positive cells in the VRNL, most probably maturing VRNs. Open arrowheads point at BrdU-positive SCs (see typical elongated nucleus)
	Fig. 2. ATP-induced [Ca2+]i increases in supporting and basal cells of the epithelium of the VNO of larval Xenopus laevis. a Pseudocolored image of an acute VNO slice stained with the Ca2+ indicator dye Fluo-4 (SCL supporting cell layer; VRNL vomeronasal receptor neuron layer; BCL basal cell layer). The upper left-hand image was acquired before application of ATP. Application of ATP-induced [Ca2+]i transients in cells of the BCL (open arrowheads) and SCL (filled arrowheads). No apparent changes in Ca2+-dependent fluorescence in cells of the VRNL (upper right-hand image). The lower left-hand image was taken after return to the base line fluorescence. A pixel correlation map (see Materials and methods for details) of the same slice is depicted in the lower right-hand image. Responsive cells appear bright on dark background. b ATP-induced [Ca2+]i transients of individual cells from the SCL (black traces). The magenta-colored area gives the mean [Ca2+]i transients ± SEM of all individual SCs (n = 20). c ATP-induced [Ca2+]i transients of individual cells from the BCL (black traces). The green-colored area gives the mean [Ca2+]i transients ± SEM of all individual BCs (n = 20)
	Fig. 3. Vomeronasal receptor neurons do not respond upon application of ATP. a A micro-Ruby backfilled and Fluo-4 stained acute slice of the vomeronasal organ (red VRNs; green Fluo-4 stained cells; SCL supporting cell layer; VRNL vomeronasal receptor neuron layer; BCL basal cell layer). b ATP and high K+-induced [Ca2+]i transients of individual micro-Ruby backfilled cells (i.e., VRNs; black traces). The red-colored areas give the mean [Ca2+]i transients ± SEM of all individual VRNs (n = 9). The VRNs did not respond upon application of ATP, but all responded upon stimulation with high K+ solution. c ATP and high K+-induced [Ca2+]i transients of individual non-neuronal cells (i.e., micro-Ruby negative cells from the SCL and BCL). The cyan-colored areas give the mean [Ca2+]i transients ± SEM of all individual non-neuronal cells (n = 11). Non-neuronal cells responded upon application of ATP, but did not respond upon stimulation with high K+
	Fig. 4. ATP-induced [Ca2+]i increases of supporting and basal cells do not depend on extracellular Ca2+ but are dependent of store depletion, and are differentially affected by the purinergic antagonist suramin. a ATP-induced [Ca2+]i increases of SCs (magenta) and BCs (green; mean response ± SEM of all ATP-responsive cells of an acute slice of the VNO; SCL supporting cell layer; BCL basal cell layer) persisted in Ca2+-free bath solution (gray-shaded rectangle; ATP application after 9 min in Ca2+-free bath solution). b Mean maximum responses ± SEM, expressed as percent of control response to ATP, of 28 SCs and 61 BCs (four VNO slices) in standard (magenta and green columns, respectively) and Ca2+-free bath solution (light magenta and light green columns, 3, 6, and 9 min in Ca2+-free bath solution, respectively). c ATP-induced [Ca2+]i increases of SCs (magenta) and BCs (green; mean response ± SEM of all ATP-responsive cells of an acute slice of the VNO) were reversibly inhibited by depletion of intracellular Ca2+ stores with CPA (10 μM, gray-shaded rectangle; ATP application after 9 min in bath solution with CPA). d Mean maximum responses ± SEM, expressed as percent of control response to ATP, of 11 SCs (two slices) and of 51 BCs (four slices) in standard (magenta and green columns, respectively) and bath solution with CPA (light magenta and light green columns, 3, 6, and 9 min in bath solution with CPA, respectively). e ATP-induced [Ca2+]i increases of SCs (magenta) and BCs (green; mean response ± SEM of all ATP-responsive cells of an acute slice of the VNO) were differentially affected by suramin (SUR, 200 μM, gray-shaded rectangle; ATP application after 9 min in bath solution with suramin). While ATP-induced [Ca2+]i increases of SCs were unaffected by suramin, the responses of BCs showed a consistent reduction in bath solution with suramin. f Mean maximum responses ± SEM, expressed as percent of control response to ATP, of 10 SCs (two slices) and of 69 BCs (seven slices) in standard (magenta and green columns, respectively) and bath solution with suramin (light magenta and light green columns, 3, 6, and 9 min in bath solution with suramin, respectively). Statistical significance was tested using paired Student’s t test (p< 0.05; *p< 0.01)
	Fig. 5. Differential responsiveness of supporting cells and basal cells to purinergic agonists. a [Ca2+]i increases of SCs in response to application of various purinergic agonists (each 100 μM; mean response ± SEM of all ATP-responsive cells of VNO slices; traces originate from more than one slice). b Mean maximum responses ± SEM, expressed as percent of control response to ATP (ATP data from 131 SCs from 14 VNO slices; ADP 47/5; adenosine 29/3; 2-MeSATP 55/6; 2-MeSADP 64/7; ATPγS 47/5; BzATP 50/6; UTP 80/8; UDP 47/5; α,β-meATP 9/2; β,γ-meATP 9/2). c [Ca2+]i increases of BCs in response to application of various purinergic agonists (each 100 μM; mean response ± SEM of all ATP-responsive cells of VNO slices; traces originate from more than one slice). d Mean maximum responses ± SEM, expressed as percent of control response to ATP (ATP data from 429 BCs from 30 VNO slices; ADP 214/14; adenosine 122/7; 2-MeSATP 163/11; 2-MeSADP 291/19; ATPγS 150/10; BzATP 105/7; UTP 189/13; UDP 188/13; α,β-meATP 81/4; and β,γ-meATP 81/4)

References [+] :

Abbracchio, Purinergic signalling in the nervous system: an overview. 2009, Pubmed