Furosemide interactions with brain GABA A receptors

• Published in: British journal of pharmacology Vol. 120; no. 5; pp. 741 - 748
• Main Authors: Korpi, Esa R, Lüddens, Hartmut
• Format: Journal Article
• Language: English
• Published: 03.02.2009
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1038/sj.bjp.0700922

The loop diuretic furosemide is known to antagonize the function of γ‐aminobutyric acid type A (GABA A ) receptors. The purpose of the present study was to examine the direct interaction of furosemide with the GABA A receptors by autoradiography and ligand binding studies with native rat and human receptors and with recombinant receptors composed of rat subunits. Autoradiography with [ 35 S]‐ t ‐butylbicyclophosphorothionate ([ 35 S]‐TBPS) as a ligand indicated that furosemide (0.1–1 m m ) reversed the 5 μ m GABA‐induced inhibition of binding only in the cerebellar granule cell layer of rat brain sections. In all other regions studied, notably also in the hippocampal and thalamic areas, furosemide failed to antagonize GABA. Furosemide 1 m m decreased [ 35 S]‐TBPS binding only in a limited number of brain regions, but facilitation of the GABA‐inhibition of the binding was much more widespread. In well‐washed rat cerebellar, but not cerebrocortical, membranes, furosemide enhanced the [ 35 S]‐TBPS binding over basal level in the absence of added GABA. The GABA A antagonist, SR 95531, and the convulsant, Ro 5–4864, blocked this furosemide‐induced increase. Both interactions with the furosemide enhancement are likely to be allosteric, since furosemide affected the binding of [ 3 H]‐SR 95531 and [ 3 H]‐Ro 5–4864 identically in the cerebellar and cerebrocortical membranes. Maximal GABA‐antagonism induced by furosemide in cerebellar membranes was further increased by SR 95531 but not by Ro 5–4864, indicating additive antagonism only for SR 95531. In human cerebellar receptors, only GABA antagonism by furosemide, but not the enhancement without added GABA, was observed. In recombinant GABA A receptors, furosemide antagonism of GABA‐inhibition of [ 35 S]‐TBPS binding depended only on the presence of α6 and β2/3 subunits, irrespective of the presence or absence of γ2 or δ subunits. In α6β3γ2 receptors, clozapine reversed the enhancement of [ 35 S]‐TBPS binding by furosemide in the absence of GABA. However, it failed to affect the GABA‐antagonism of furosemide, suggesting that the enhancement of basal binding and the GABA antagonism might represent two different allosteric actions of furosemide. In conclusion, the present results indicate that furosemide is a subtype‐selective GABA A antagonist with a mode of action not shared by several other antagonists, which makes furosemide a unique compound for development of potential GABA A receptor subtype‐specific and ‐selective ligands. British Journal of Pharmacology (1997) 120 , 741–748; doi: 10.1038/sj.bjp.0700922