Purinergic calcium responses in murine subcutaneous white fat.

P. Smith. Life Sciences, University of Nottingham, Nottingham, United Kingdom.

Introduction: Sympathetic innervation of white adipose tissue mediate cellular responses by noradrenaline and the co-transmitter ATP via action at beta-adrenoceptors and purinergic receptors respectively. Canonically, ATP mobilises intracellular calcium via P2Y2 receptor activation, the aim was to determine whether this also occurs in subcutaneous white fat adipose tissue (SCWAT) (1).

Methods: Intracellular calcium, [Ca²⁺], was imaged using epifluorescent microscopy of adipocytes harvested from the SCWAT of 10 male CD1 mice following published procedures (2, 3). Data are reported as medians and interquartile range. Results SCWAT had a basal [Ca²⁺] of 117 nM (62-184; n = 137). With 100 μM ATP, 79% of cells gave a transient uniform increase in [Ca²⁺] which peaked at 384 nM (226-845; n = 78); of these 62% monotonically decayed back to basal whereas 37% possessed a maintained plateau; however only the latter ever gave a response to a 2^nd application of ATP with an infinite odds ratio. In the 33% of cells that failed to respond to a 2nd application of ATP, a transient increase in [Ca²⁺] could be evoked by 1 μM oxytocin, this peaked at a similar value to that of the 1^st ATP response: 335 nM (270-576; n = 20), data indicative of the existence of a functional Ca²⁺ mobilization pathway. Virgin application of oxytocin gave a uniform [Ca²⁺] increase in 86% of adipocytes, which peaked at 685 nM (404-3973; n = 20); of these 66% monotonically decayed back to basal whereas 34% had a plateau; subsequent application of ATP transiently increased [Ca²⁺] in 90% of cells tested regardless of an oxytocin-evoked [Ca²⁺] plateau. Virgin application of 100 μM UTP, a P2Y2 agonist, evoked a transient increase in [Ca²⁺] which peaked at 370 nM (262-724; n = 10) of which all subsequently responded to ATP independent of a UTP plateau. 2-APB, an inhibitor of store operated calcium release did not affect the ATP response.

Conclusions: SCWAT has a varied distribution of P2Y2 receptors, whose activation mobilizes intracellular Ca²⁺ with heterogeneous responses. Interestingly the presence of a plateau appears unrelated to store refilling but to receptor kinetics.

References:

1. Lee H et al (2005) J Biol Chem 280: 28556-28

2. Bentley D et al (2014). Am..J. Physiol. 307: E161-E175.

3. Komai AM et al (2016) Diabetes 65: 3301-3313.