Stuart C. Mangel - US grants

Although the receptive field properties of mammalian ganglion cells are quite pertinent to visual function, little is known concerning the presynaptic pathways which underlie the properties of these cells. This project proposes to examine the influence of horizontal cells upon the receptive field properties of ganglion cells the rabbit retina by the use of intracellular and extracellular recording techniques and by pharmacological manipulations. In particular, horizontal cells will be artificially polarized by intracellular current injections and the resultant effects upon visually-evoked ganglion cell activity studied. These experiments will determine whether horizontal cells mediate the surround responses of ganglion cells in the rabbit retina and whether other ganglion cell response properties, such as spatial resolution and orientation selectivity, are also influenced by horizontal cells. In addition, because different types of rabbit horizontal cells receive different proportions of rod and cone input, horizontal cell polarization of these cell types will reveal whether rod and cone pathways exert differential effects upon ganglion cell receptive field properties. Finally, the retinal pathways between horizontal and ganglion cells will be further characterized by the application of neurotransmitter blocking agents to determine whether these drugs mimic or block the effects of horizontal cell polarization upon particular properties of ganglion cells. Knowledge of neural circuitry in a mammalian retina will increase understanding of human retinal processes and enhance the diagnosis of retinal pathologies. These data from the rabbit retina will also increase understanding of the ERG and thus will enhance the diagnostic value of the ERG and improve its usefulness as a tool in ophthalmological research.

DESCRIPTION (Adapted from applicant's abstract): This project seeks to understand how neuronal circuits in the rabbit retina change or adapt due to a circadian oscillator. A circadian clock or oscillator has a rhythmicity of approximately 24 hours in the absence of external timing cues. Cyclic environmental stimuli such as light can entrain the circadian clock. This project will determine whether a circadian clock regulates cone-horizontal cells that exhibit a diurnal rhythm. The dominant photoreceptor input to dark-adapted, cone horizontal cells exhibits a diurnal rhythm; cone inputs predominate during the day and rod inputs predominate during the night. A combination of electrophysiological, neurochemical and anatomical techniques will be utilized to determine whether a circadian clock is involved in horizontal cell and ganglion cell light responsiveness, as well as horizontal cell gap junctional coupling. It will be determined whether a circadian clock controls the levels of dopamine, melatonin and adenosine, as well as extracellular pH. It will then be determined whether the circadian clock uses any of these signaling mechanisms to regulate the light responses of horizontal and ganglion cells. Measurements will be made in constant darkness (absence of timing cues) in a superfused, retinal eyecup preparation. Disruption of the circadian clock may result in retinal photoreceptor degeneration. Thus, increased understanding of the circadian clock processes will aid in the understanding of human retinal processes and dysfunction. These studies may also provide the basis for drug therapy for retinal disorders thought to be linked to the disruption of circadian processes.

[unreadable] DESCRIPTION (provided by applicant): This research project is an experimental study that seeks to understand the role of cation-chloride co-transporters in information processing in the mammalian retina. In the retina and elsewhere in the central nervous system, two types of chloride co-transporters, the Na-K-Cl and K-Cl co-transporters, have been identified. These chloride co-transporters regulate the intracellular chloride concentration such that the K-Cl co-transporter extrudes chloride from neurons, whereas the Na-K-Cl co-transporter transports chloride into cells. Thus, the neurotransmitter GABA hyperpolarizes neurons when the chloride equilibrium potential is less than the resting membrane potential due to the action of the K-Cl co-transporter. In contrast, GABA depolarizes neurons when the chloride equilibrium potential is greater than the resting membrane potential due to the action of the Na-K-Cl co-transporter. Thus, depending on the type of chloride co-transporter expressed by a retinal neuron, GABA will either hyperpolarize or depolarize the cell.We will therefore determine the roles of chloride co-transporters in the retina by using a combination of electrophysiological, neurochemical and anatomical techniques. A rabbit eyecup preparation will be used to study the roles of chloride co-transporters in directional selectivity and in synaptic transmission from horizontal cells to bipolar cells. Specifically, we will determine whether an asymmetric distribution of the Na-K-Cl and K-Cl co-transporters on starburst amacrine cells mediates the null direction inhibition exhibited by rabbit ON-OFF directionally-selective ganglion cells. We will also determine whether horizontal cells contribute to the receptive field surround of ON-center bipolar cells (and ON-center ganglion cells) in part by a direct, GABA-mediated synaptic connection that is Na-K-Cl co-transporter-dependent and sign conserving. Finally, we will determine whether and how the development of chloride co-transporter function affects these retinal phenomena.Increased knowledge of chloride co-transporter function in the adult and developing retina will aid in the understanding of human retinal processes and dysfunction, as well as provide the basis for drug therapy for retinal disorders.

This research project is an experimental study that seeks to understand how neuronal networks change or adapt due to the response of the retina to the gradual change in the ambient light level that occurs day and night, and the influence of the circadian (24-h) clock that is intrinsic to the retina. The release of the neuromodulator dopamine in the retina is controlled by the retinal clock, which increases dopamine levels sufficiently at dawn to activate the highly sensitive dopamine D4 receptors on cones. In addition, distinct non- circadian light responsive processes increase dopamine levels to a much greater extent in response to bright illumination at midday so that the less sensitive dopamine D1 receptors on dendrites of cone bipolar cells, a type of second order cell that receives synaptic input from cones, are activated. The proposed experiments will study whether the bright light-induced increase in D1 receptor activation strengthens GABA signaling from horizontal cells (another type of second order cell that receives cone input) to cone bipolar cells in the day by enhancing GABAA receptor function of cone bipolar cell dendrites. The proposed experiments will also investigate whether the retinal clock, by decreasing D4 receptor activation at night, strengthens GABA signaling from horizontal cells to cones at night in the dark by enhancing GABAA receptor function of cone synaptic terminals. Electrophysiological recording in rabbit retinal slices will be used to study the light responses and GABAA receptor activity of cone bipolar cells and GABA signaling from horizontal cells to cone bipolar cells. Also, electrophysiological recording in intact goldfish and rabbit neural retinas will be used to study the light responses and GABAA receptor activity of cones and GABA signaling from horizontal cells to cones. Neurochemical, cell/molecular, and anatomical techniques will also be employed using intact rabbit and fish retinas, studied in the day and night under constant darkness, and in the day following maintained illumination.