Up-Regulation of Excitatory Amino Acid Transporters EAAT1 and EAAT2 by ß-Klotho

Abstract

Background/Aims: Klotho, a transmembrane protein expressed in chorioid plexus of the brain, kidney, and several other tissues, is required for inhibition of 1,25(OH)₂D₃ formation by FGF23. The extracellular domain of Klotho protein could be cleaved off, thus being released into blood or cerebrospinal fluid. At least in part by exerting β-glucuronidase activity, soluble klotho regulates several ion channels and carriers. Klotho protein deficiency accelerates the appearance of age related disorders including neurodegeneration and muscle wasting and eventually leads to premature death. The present study explored the effect of Klotho protein on the excitatory glutamate transporters EAAT1 (SLC1A3) and EAAT2 (SLC1A2), Na⁺ coupled carriers clearing excitatory amino acids from the synaptic cleft and thus participating in the regulation of neuronal excitability. Methods: cRNA encoding EAAT1 or EAAT2 was injected into Xenopus laevis oocytes and glutamate (2 mM)-induced inward current (I_Glu) taken as measure of glutamate transport. Measurements were made without or with prior 24 h treatment with soluble ß-Klotho protein (30 ng/ml) in the absence and presence of β-glucuronidase inhibitor D-saccharic acid 1,4-lactone monohydrate (DSAL,10 µM). Results: I_Glu was observed in EAAT1 and in EAAT2 expressing oocytes but not in water injected oocytes. In both, EAAT1 and EAAT2 expressing oocytes I_Glu was significantly increased by treatment with soluble ß-Klotho protein, an effect reversed by DSAL. Treatment with ß-klotho protein increased significantly the maximal transport rate without significantly modifying the affinity of the carriers. Conclusion: ß-Klotho up-regulates the excitatory glutamate transporters EAAT1 and EAAT2 and thus participates in the regulation of neuronal excitation.

β-Klotho as a Negative Regulator of the Peptide Transporters PEPT1 and PEPT2

“…Klotho deficiency is followed by excessive 1,25(OH) 2 D 3 formation [2,13,14] leading to increase of plasma Ca 2+ [18] and phosphate [17] concentrations as well as vascular calcification [19]. Besides its effect on 1,25(OH) 2 D 3 formation, klotho modifies the function of several transport proteins including Ca 2+ channels [20], Na + ,phosphate cotransport [4,21], Na + /K + ATPase [22], renal outer medullary K + channels [23], Kv1.3 K + channels [24], KCNQ1/KCNE1 [25], the creatine transporter CreaT [26], and the excitatory amino acid transporters EAAT1, EAAT2, EAAT3 and EAAT4 [27,28].…”

SGK3 Sensitivity of Voltage Gated K+ Channel Kv1.5 (KCNA5)

“…Constructs encoding mouse wild-type KCNA5 [23,24], wild type SGK3 [25], constitutively active S419D SGK3 [26], inactive K191N SGK3 [26] and wild type Nedd4-2 [2] were used for generation of cRNA as described previously [27][28][29].…”

Up-Regulation of the Excitatory Amino Acid Transporters EAAT1 and EAAT2 by Mammalian Target of Rapamycin

“…The constructs were sequenced to verify the correctness of the plasmid [50]. The constructs were used for generation of cRNA as described previously [51][52][53].…”