Reciprocal regulation between taurine and glutamate response via Ca2+-dependent pathways in retinal third-order neurons

Abstract

Although taurine and glutamate are the most abundant amino acids conducting neural signals in the central nervous system, the communication between these two neurotransmitters is largely unknown. This study explores the interaction of taurine and glutamate in the retinal third-order neurons. Using specific antibodies, both taurine and taurine transporters were localized in photoreceptors and Off-bipolar cells, glutamatergic neurons in retinas. It is possible that Off-bipolar cells release juxtaposed glutamate and taurine to activate the third-order neurons in retina. The interaction of taurine and glutamate was studied in acutely dissociated third-order neurons in whole-cell patch-clamp recording and Ca2+ imaging. We find that taurine effectively reduces glutamate-induced Ca2+ influx via ionotropic glutamate receptors and voltage-dependent Ca2+ channels in the neurons, and the effect of taurine was selectively inhibited by strychnine and picrotoxin, but not GABA receptor antagonists, although GABA receptors are present in the neurons. A CaMKII inhibitor partially reversed the effect of taurine, suggesting that a Ca2+/calmodulin-dependent pathway is involved in taurine regulation. On the other hand, a rapid influx of Ca2+ through ionotropic glutamate receptors could inhibit the amplitude and kinetics of taurine-elicited currents in the third-order neurons, which could be controlled with intracellular application of BAPTA a fast Ca2+ chelator. This study indicates that taurine is a potential neuromodulator in glutamate transmission. The reciprocal inhibition between taurine and glutamate in the postsynaptic neurons contributes to computation of visual signals in the retinal neurons.

Figure 1

Taurinergic neurons and taurine–dose response in salamander retina. Immunoreactive pattern shows that taurine labeling is present in rods, a few cones and displaced Off-bipolar cells (asterisks) in the ONL. It is also present in cell somas in the INL and axon terminals in the sublamina of the IPL (A); anti-taurine transporter labels rod axons and Off-bipolar cell axons in the sublamina a of the IPL (B). Whole-cell recording of taurine dose response currents from an isolated ganglion cell (C). Taurine dose-response curve and EC₅₀ value were obtained from the third-order neurons.

Figure 2

Statistical results of taurine regulation of glutamate-induce [Ca²⁺]_i in isolated third-order neurons. Taurine suppresses the [Ca²⁺]_i in the third-order neurons in a dose- dependent manner (A). Suppressive effects of taurine on AMPA-, kainate- and NMDA-induced [Ca²⁺]_i in the neurons (B, C and D).

Figure 3

Pharmacological study of taurine response in isolated third-order neurons. Strychnine (2µM) partially reverses the suppressive effects of taurine on glutamate-, AMPA- and kainate-induced [Ca²⁺]_i from the third-order neurons (A, B and C). Picrotoxin (100µM) had similar effects as strychnine partially reversing the effects of taurine on glutamate- and AMPA-induced [Ca²⁺]_i (D and E).

Figure 4

GABA receptor antagonists had no effect on taurine-produced effects. Neither GABA_A receptor antagonists, bicuculline (10µM) and SR95531 (10µM) (A and B), nor GABA_C and GABA_B receptor antagonists, TPMPA (50µM) and CGP55845 (10µM; C and D) block taurine suppression on glutamate-induced [Ca²⁺]_i .

Figure 5

Glutamate induces Ca²⁺ influx via both glutamate receptors and voltage-gated Ca²⁺ channels in third-order neurons. (A) Glutamate-induced [Ca²⁺]_i is completely blocked by ionotropic glutamate receptor antagonists, CNQX (50µM) and AP-7(40µM). (B) Co²⁺ blocks voltage-gated Ca²⁺ channels, which reduces 55% of glutamate-induced [Ca²⁺]_i in the neurons (see asterisk); with Co²⁺ taurine still suppresses glutamate-induced [Ca²⁺]_i. (C) Ruthenium red (1µM), a rynadine receptor inhibitor, reduces 15% of glutamate-induced [Ca²⁺]_i; with ruthenium red, taurine has 35% suppression on glutamate-induced [Ca²⁺]_i in the neurons. (D) Blocking Ca²⁺ uptake pumps in the internal organelles with thapsigargin (1µM) had a limited effect on taurine regulation of [Ca²⁺]_i.

Figure 6

Pharmacological evidence of CaMKII and PKA involved in taurine regulation of glutamate-induced [Ca²⁺]_i . (A) Statistical results of the selective cell-permeable CaM kinase II inhibitor KN-62 and protein kinase A inhibitor PKI (14-22) amide (both 1µM) on taurine regulation of glutamate-induced [Ca²⁺]_i in isolated third-order neurons. (B) CaMKII involved in the strychnine-insensitive taurine regulation on glutamate response.

Figure 7

Taurine-elicited currents are suppressed by AMPA and kainate in isolated third-order neurons. (A and B) 350µM taurine elicits currents that are suppressed by co-application of either 15µM AMPA or kainate; both amplitudes and kinetics of the taurine currents are altered in the presence of AMPA or kainate. (C and D) the same experiments repeated with intracellular application of 10mM BAPTA.

Figure 8

Potential intracellular pathways for reciprocal regulations between taurine-sensitive receptor and glutamate receptor.