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Figure 2 | BMC Pharmacology

Figure 2

From: Lipid phosphate phosphatase inhibitors locally amplify lysophosphatidic acid LPA1 receptor signalling in rat brain cryosections without affecting global LPA degradation

Figure 2

Vanadate and propranolol evoke LPA-mimicking [35S]GTPγS binding response that is sensitive to LPA 1/3 receptor antagonist. (a) Coronal brain sections were incubated using a three-step autoradiography protocol as detailed in Methods. Test chemicals were included during the [35 S]GTPγS labelling step (step 3) in the presence of 0.1% BSA. Treatment with propranolol (1 mM) or Na3VO4 (100 μM), results in G protein activity in the LPA1 receptor enriched white matter tracts, a response that is mimicked by exogenous LPA (50 μM). The LPA1/3 receptor antagonist Ki16425 (5 μM) abolishes all the evoked responses in the white matter regions, including the tonic LPA1 signal observed under basal conditions (cc, corpus callosum). Scale bar = 2 mm. The eight-point [14 C] standard used in the quantification is shown at the bottom of image. (b) and (c) Quantitative data of the binding responses including the dose response for exogenous LPA (0.5 μM to 50 μM) with or without the simultaneous treatment with the inhibitors. Autoradiography films were digitized and [35 S]GTPγS binding was quantified from the corpus callosum of coronal sections of 4 week-old rat brain, as described in Methods. Quantitative data are calculated as nCi/g equivalents with non-specific binding subtracted from total binding. The data are expressed as a percentage of basal binding (mean + SEM) from six individual animals (n = 6). Significance level: (b) ***p < 0.001 compared to the treatment without Ki16425 (c) ***p < 0.001 compared to control in each LPA concentration.

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