Intracellular recordings were obtained from rods in the Gekko gekko retina and the adaptation characteristics of their responses studied during light and dark adaptation. Steady background illumination induced graded and sustained hyperpolarizing potentials and compressed the incremental voltage range of the receptor. Steady backgrounds also shifted the receptor's voltageintensity curve along the intensity axis, and bright backgrounds lowered the saturation potential of the receptor. Increment thresholds of single receptors followed Weber's law over a range of about 3.5 log units and then saturated. Most of the receptor sensitivity change in light derived from the shift of the voltage-intensity curve, only little from the voltage compression. Treatment of the eyecup with sodium aspartate at concentrations sufficient to eliminate the b-wave of the eleetroretinogram (ERG) abolished initial transients in the receptor response, possibly indicating the removal of horizontal cell feedback. Aspartate treatment, however, did not significantly alter the adaptation characteristics of receptor responses, indicating that they derive from processes intrinsic to the receptors. Dark adaptation after a strongly adapting stimulus was similarly associated with temporary elevation of membrane potential, initial lowering of the saturation potential, and shift of the voltage-intensity curve. Under all conditions of adaptation studied, small amplitude responses were linear with light intensity. Further, there was no unique relation between sensitivity and membrane potential suggesting that receptor sensitivity is controlled at least in part by a step of visual transduetion preceding the generation of membrane voltage change.

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We tested the effect of anions on the absorbance spectrum of native visual pigments as measured by microspectrophotometry in individual cone outer segments of four species of fish and one species of amphibian. In all species tested, the long-wavelength-absorbing cone pigments were anion sensitive, and their A.. could be tuned over a range of 55 nm depending on the identity of the anion present. Cl-and Br-were the only anions that produced native pigment spectra by red shifting A.. from its value under anion-free conditions. Lyotropic anions such as NO-, SCN-, BF4, and ClIO caused substantial and graded blue shifts of A... The apparent Kd of binding sites on the pigment for Cl-and for CIO4 was -2 mM. Taken together with previous findings on three visual pigments from the reptilian, avian, and amphibian classes, our results support the hypothesis that all long-wavelength-absorbing vertebrate visual pigments are spectrally tuned in part through the binding of a chloride ion. We propose that the site of anion tuning is near the protonated Schiff base of the chromophore, whose counterion may be complex and include Cl-as an exchangeable anion. This counterion configuration may resemble the one present in the light-driven Cl-pump halorhodopsin.Color vision in vertebrates is based on sets of two, three, or four different photopigments that reside in separate classes of cone photoreceptors. The wavelength of peak absorbance (Amax) of these pigments ranges from the UV (360 nm) to the far red (635 nm). All vertebrate visual pigments are integral membrane proteins and contain 11-cis-retinal or 11-cisdehydroretinal as the chromophore that is covalently bound through a Schiffbase linkage to a lysine residue on the protein (opsin) moiety ofthe pigment (1). Although great progress has been made in recent years in unraveling the structure of visual pigments, the molecular basis of spectral tuning is still only incompletely understood.A variety of mechanisms have been invoked to account for photopigment tuning (for reviews, see refs. 2 and 3). For example, protonation of the retinal Schiff base shifts Amax from 360 to 430 nm (4). Blue-absorbing pigments may contain an unperturbed chromophore behaving much like a protonated retinal Schiff base in a nonpolar solvent (5). Closer interaction of the chromophore with the protein perturbs the electronic structure of the chromophore and results in an additional red shift of Am,,,. This shift is commonly called the "opsin shift" and very likely comprises multiple components. For example, Amax is extremely sensitive to the charge environment provided by the counterion, which is paired with the protonated Schiff base (3). A decreased interaction between protonated Schiff base and counterion, perhaps caused by an increased distance between the two, may be responsible for the opsin shift in green-absorbing rhodopsins (Amax, "500 nm). Amino acid side chains, which form the hydrophobic binding pocket for the chromophore, also can interact with and perturb the chromophore. In fact, much ...

We present microspectrophotometric evidence for the existence of two distinct visual pigments residing in two different morphological types of photoreceptor of the sea lamprey. In the upstream migrant Petromyzon marinus, the pigment found in short receptors has a wavelength of peak absorbance (λmax) of 525 nm, whereas the pigment located in long receptors has a λmax of 600 nm. Although the former appears to be pure porphyropsin, the latter is akin to visual pigments found in the red-absorbing cones of amphibian and teleost retinae. The kinship is more than superficial pertaining to λmax of the a–band absorbance to its native maximum value. The presence of an anion-sensitive and an anion-insensitive pigment in a retina implies the expression of two distinct opsin genes. We infer this from several examples of correlation between anion sensitivity and opsin sequence groupings. Moreover, the presence of two distinct opsin genes expressed throughout six vertebrate classes implies their existence in a common ancestor to all.