cGMP modulates spike responses of retinal ganglion cells via a cGMP-gated current FUSAO KAWAI 1,2 and PETER STERLING 1 1 Department of Neuroscience, University of Pennsylvania, Philadelphia 2 Department of Physiology, Fujita Health University, Toyoake, Aichi, 470-1192, Japan (Received December 12, 2001; Accepted April 25, 2002) Abstract Certain ganglion cells in the mammalian retina are known to express a cGMP-gated cation channel. We found that a cGMP-gated current modulates spike responses of the ganglion cells in mammalian retinal slice preparation. In such cells under current clamp, bath application of the membrane-permeant cGMP analog (8-bromo-cGMP, 8-p-chlorophenylthio-cGMP) or a nitric oxide donor (sodium nitroprusside, S-nitroso-N-acetyl-penicillamine) depolarized the membrane potential by 5–15 mV, and reduced the amount of current needed to evoke action potentials. Similar effects were observed when the membrane potential was simply depolarized by steady current. The responses to cGMP are unaffected by inhibitors of cGMP-dependent protein kinase and Ca 2+ 0calmodulin-dependent protein kinase. The response to cGMP persisted in Ca 2+ -free bath solution with Ca 2+ buffers in the pipette. Under voltage clamp, cGMP analogs did not affect the response kinetics of voltage-gated currents. We conclude that cGMP modulates ganglion cell spiking simply by depolarizing the membrane potential via the inward current through the cGMP-gated channel. Modulation of this channel via the long-range NO-synthase amacrine cell may contribute to control of contrast gain by peripheral mechanisms. Keywords: Retina, Ganglion cell, Guinea pig, Patch clamp, cGMP-gated channel Introduction Certain ganglion cells in mammalian retina express a cGMP-gated cation channel (Ahmad et al., 1994; Kawai & Sterling, 1999). This channel is modulated when nitric oxide (NO) released from am- acrine cells (Sandell, 1985; Vincent & Kimura, 1992; Haberecht et al., 1998) stimulates the ganglion cell’s soluble guanylyl cyclase (Ahmad & Barnstable 1993; Gotzes et al., 1998; Kawai & Sterling, 1999). The effect of current through this channel had not been studied and was complicated to address because cGMP can have several effects. For example, cGMP can modulate voltage-gated currents directly (Levitan & Levitan, 1988) and via cGMP- dependent protein kinase (PKG) (Paupardin-Tritsch et al., 1986; Sumii & Sperelakis, 1995; Lohmann et al., 1997; Wei et al., 1998). Here we investigated the effect of current through the cGMP-gated channel on ganglion cell responses, isolating this effect from that of the PKG cascade and from the effect of Ca 2+ entering via the cGMP-gated channel on voltage-gated currents. Methods Preparation and recording Slices from adult guinea pig retina were cut at 200 mm (Kawai & Sterling, 1999) and viewed on a Zeiss upright microscope with differential interference contrast optics (40 water-immersion objective). Ganglion cells were identified in the slice by their position and size. Membrane voltages and currents were recorded in the whole-cell configuration using a patch-clamp amplifier (Axopatch 200A, Axon Instruments, Foster City, CA) linked to a computer. The current- and voltage-clamp procedures were con- trolled by the pCLAMP software (Axon Instruments). Data were low-pass filtered (4-pole Bessel type) with a cut-off frequency of 5 kHz and then digitized at 10 kHz by an analog-to-digital interface. All experiments were performed at room temperature (23–258C). Solutions and drugs In most experiments, the tissue was perfused with Ames medium (which is buffered with 22.6 mM bicarbonate), bubbled with 95% O 2 05% CO 2 . In some experiments, Ringer’s solution was used instead of Ames’ solution to eliminate Ca 2+ in the bath. The Ringer’s solution contained (in mM) 135 NaCl; 5 KCl; 10 HEPES; Address correspondence and reprint requests to: Fusao Kawai, Depart- ment of Physiology, School of Medicine, Fujita Health University, 1-98 Dengakugakubo, Kutsukakechou, Toyoake, Aichi, 470-1192, Japan. E-mail: fkawai@fujita-hu.ac.jp Visual Neuroscience (2002), 19, 373–380. Printed in the USA. Copyright © 2002 Cambridge University Press 0952-5238002 $12.50 DOI: 10.1017.S0952523802193138 373