Starburst Cells Research Articles

Two mechanisms for direction selectivity in a model of the primate starburst amacrine cell.

In a recent study, visual signals were recorded for the first time in starburst amacrine cells of the macaque retina, and, as for mouse and rabbit, a directional bias observed in calcium signals was recorded from near the dendritic tips. Stimulus motion from the soma toward the tip generated a larger calcium signal than motion from the tip toward the soma. Two mechanisms affecting the spatiotemporal summation of excitatory postsynaptic currents have been proposed to contribute to directional signaling at the dendritic tips of starbursts: (1) a "morphological" mechanism in which electrotonic propagation of excitatory synaptic currents along a dendrite sums bipolar cell inputs at the dendritic tip preferentially for stimulus motion in the centrifugal direction; (2) a "space-time" mechanism that relies on differences in the time-courses of proximal and distal bipolar cell inputs to favor centrifugal stimulus motion. To explore the contributions of these two mechanisms in the primate, we developed a realistic computational model based on connectomic reconstruction of a macaque starburst cell and the distribution of its synaptic inputs from sustained and transient bipolar cell types. Our model suggests that both mechanisms can initiate direction selectivity in starburst dendrites, but their contributions differ depending on the spatiotemporal properties of the stimulus. Specifically, the morphological mechanism dominates when small visual objects are moving at high velocities, and the space-time mechanism contributes most for large visual objects moving at low velocities.

Impairment of amacrine starburst cells and their synaptic contacts with dopaminergic cells could explain motion perception disturbances in Parkinson's disease

AbstractPurpose: Parkinson's disease (PD) patients suffer from visual deficiencies in the early stages of the disease. One of these visual symptoms is the impairment of the motion perception. In that sense, we wanted to determine if the starburst amacrine cells (the main cell type involved in motion perception) are degenerated in PD. Also, we sought to evaluate if this possible degeneration is related to the degeneration of the dopaminergic system.Methods: Post‐mortem human eyes from control and PD donors were used in this study. Immunohistochemistry and confocal microscopy were used in whole‐mount retinas to evaluate tyrosine hydroxylase‐positive cell density (dopaminergic amacrine cells), and choline acetyltransferase‐positive cell density (starburst amacrine cell). Moreover, we quantified the dopaminergic synaptic contacts with starburst amacrine cells using vesicular monoamine transporter‐2 (VMAT2) antibodies.Results: There is a decrease in the dopaminergic amacrine cell density in PD retinas. Furthermore, we found a decrease in the number of starburst amacrine cells. Importantly, we describe for the first time that dopaminergic amacrine cells contact with starburst amacrine cells in the retina and that these connections decrease in PD patients.Conclusions: This work shows the degeneration of starburst amacrine cells and their synaptic connections with dopaminergic cells, and this may explain the disturbances in motion perception present in PD. Importantly, these deficiencies can be used as a biomarker of this pathology performing functional tests for visual motion perception.Support: Ministerio de Ciencia e Innovación (FEDER‐ PID2019‐106230RB‐I00). Ministerio de Universidades (FPU16/04114). Instituto Carlos III (RETICS‐FEDER RD16/0008/0016). Generalitat Valenciana (IDIFEDER/2017/064, ACIF/2020/203). Michael J Fox Foundation for Parkinson's Research.

Origins of direction selectivity in the primate retina

From mouse to primate, there is a striking discontinuity in our current understanding of the neural coding of motion direction. In non-primate mammals, directionally selective cell types and circuits are a signature feature of the retina, situated at the earliest stage of the visual process. In primates, by contrast, direction selectivity is a hallmark of motion processing areas in visual cortex, but has not been found in the retina, despite significant effort. Here we combined functional recordings of light-evoked responses and connectomic reconstruction to identify diverse direction-selective cell types in the macaque monkey retina with distinctive physiological properties and synaptic motifs. This circuitry includes an ON-OFF ganglion cell type, a spiking, ON-OFF polyaxonal amacrine cell and the starburst amacrine cell, all of which show direction selectivity. Moreover, we discovered that macaque starburst cells possess a strong, non-GABAergic, antagonistic surround mediated by input from excitatory bipolar cells that is critical for the generation of radial motion sensitivity in these cells. Our findings open a door to investigation of a precortical circuitry that computes motion direction in the primate visual system.

Direction selectivity in retinal bipolar cell axon terminals

SummaryThe ability to encode the direction of image motion is fundamental to our sense of vision. Direction selectivity along the four cardinal directions is thought to originate in direction-selective ganglion cells (DSGCs) because of directionally tuned GABAergic suppression by starburst cells. Here, by utilizing two-photon glutamate imaging to measure synaptic release, we reveal that direction selectivity along all four directions arises earlier than expected at bipolar cell outputs. Individual bipolar cells contained four distinct populations of axon terminal boutons with different preferred directions. We further show that this bouton-specific tuning relies on cholinergic excitation from starburst cells and GABAergic inhibition from wide-field amacrine cells. DSGCs received both tuned directionally aligned inputs and untuned inputs from among heterogeneously tuned glutamatergic bouton populations. Thus, directional tuning in the excitatory visual pathway is incrementally refined at the bipolar cell axon terminals and their recipient DSGC dendrites by two different neurotransmitters co-released from starburst cells.

Bistratified starburst amacrine cells in Sox2 conditional knockout mouse retina display ON and OFF responses.

Cell-intrinsic factors, in conjunction with environmental signals, guide migration, differentiation, and connectivity during early development of neuronal circuits. Within the retina, inhibitory starburst amacrine cells (SBACs) comprise ON types with somas in the ganglion cell layer (GCL) and dendrites stratifying narrowly in the inner half of the inner plexiform layer (IPL) and OFF types with somas in the inner nuclear layer (INL) and dendrites stratifying narrowly in the outer half of the IPL. The transcription factor Sox2 is crucial to this subtype specification. Without Sox2, many ON-type SBACs destined for the GCL settle in the INL while many that reach the GCL develop bistratified dendritic arbors. This study asked whether ON-type SBACs in Sox2-conditional knockout retinas exhibit selective connectivity only with ON-type bipolar cells or their bistratified morphology allows them to connect to both ON and OFF bipolar cells. Physiological data demonstrate that these cells receive ON and OFF excitatory inputs, indicating that the ectopically stratified dendrites make functional synapses with bipolar cells. The excitatory inputs were smaller and more transient in Sox2-conditional knockout compared with wild type; however, inhibitory inputs appeared largely unchanged. Thus dendritic stratification, rather than cellular identification, may be the major factor that determines ON vs. OFF connectivity. NEW & NOTEWORTHY Conditional knockout of the transcription factor Sox2 during early embryogenesis converts a monostratifying starburst amacrine cell into a bistratifying starburst cell. Here we show that these bistratifying starburst amacrine cells form functional synaptic connections with both ON and OFF bipolar cells. This suggests that normal ON vs. OFF starburst connectivity may not require distinct molecular specification. Proximity alone may be sufficient to allow formation of functional synapses.

Synaptic connections of amacrine cells containing vesicular glutamate transporter 3 in baboon retinas.

The goals of these experiments were to describe the morphology and synaptic connections of amacrine cells in the baboon retina that contain immunoreactive vesicular glutamate transporter 3 (vGluT3). These amacrine cells had the morphology characteristic of knotty bistratified type 1 cells, and their dendrites formed two plexuses on either side of the center of the inner plexiform layer. The primary dendrites received large synapses from amacrine cells, and the higher-order dendrites were both pre- and postsynaptic to other amacrine cells. Based on light microscopic immunolabeling results, these include AII cells and starburst cells, but not the polyaxonal amacrine cells tracer-coupled to ON parasol ganglion cells. The vGluT3 cells received input from ON bipolar cells at ribbon synapses and made synapses onto OFF bipolar cells, including the diffuse DB3a type. Many synapses from vGluT3 cells onto retinal ganglion cells were observed in both plexuses. At synapses where vGluT3 cells were presynaptic, two types of postsynaptic densities were observed; there were relatively thin ones characteristic of inhibitory synapses and relatively thick ones characteristic of excitatory synapses. In the light microscopic experiments with Neurobiotin-injected ganglion cells, vGluT3 cells made contacts with midget and parasol ganglion cells, including both ON and OFF types. Puncta containing immunoreactive gephyrin, an inhibitory synapse marker, were found at appositions between vGluT3 cells and each of the four types of labeled ganglion cells. The vGluT3 cells did not have detectable levels of immunoreactive γ-aminobutyric acid (GABA) or immunoreactive glycine transporter 1. Thus, the vGluT3 cells would be expected to have ON responses to light and make synapses onto neurons in both the ON and the OFF pathways. Taken with previous results, these findings suggest that vGluT3 cells release glycine at some of their output synapses and glutamate at others.

Spatially asymmetric reorganization of inhibition establishes a motion-sensitive circuit

Spatial asymmetries in neural connectivity have an important role in creating basic building blocks of neuronal processing. A key circuit module of directionally selective (DS) retinal ganglion cells is a spatially asymmetric inhibitory input from starburst amacrine cells. It is not known how and when this circuit asymmetry is established during development. Here we photostimulate mouse starburst cells targeted with channelrhodopsin-2 (refs 6-8) while recording from a single genetically labelled type of DS cell. We follow the spatial distribution of synaptic strengths between starburst and DS cells during early postnatal development before these neurons can respond to a physiological light stimulus, and confirm connectivity by monosynaptically restricted trans-synaptic rabies viral tracing. We show that asymmetry develops rapidly over a 2-day period through an intermediate state in which random or symmetric synaptic connections have been established. The development of asymmetry involves the spatially selective reorganization of inhibitory synaptic inputs. Intriguingly, the spatial distribution of excitatory synaptic inputs from starburst cells is significantly more symmetric than that of the inhibitory inputs at the end of this developmental period. Our work demonstrates a rapid developmental switch from a symmetric to asymmetric input distribution for inhibition in the neural circuit of a principal cell.

Synaptic Regulation of the Light-Dependent Oscillatory Currents in Starburst Amacrine Cells of the Mouse Retina

Responses of on-center starburst amacrine cells to steady light stimuli were recorded in the dark-adapted mouse retina. The response to spots of dim white light appear to show two components, an initial peak that correspond to the onset of the light stimulus and a series of oscillations that ride on top of the initial peak relaxation. The frequency of oscillations during light stimulation was three time higher than the frequency of spontaneous oscillations recorded in the dark. The light-evoked responses in starburst cells were exclusively dependent on the release of glutamate likely from presynaptic bipolar axon terminals and the binding of glutamate to AMPA/kainate receptors because they were blocked by 6-cyano-7-nitroquinoxalene-2,3-dione. The synaptic pathway responsible for the light responses was blocked by AP4, an agonist of metabotropic glutamate receptors that hyperpolarize on-center bipolar cells on activation. Light responses were inhibited by the calcium channel blockers cadmium ions and nifedipine, suggesting that the release of glutamate was calcium dependent. The oscillatory component of the response was specifically inhibited by blocking the glutamate transporter with d-threo-beta-benzyloxyaspartic acid, suggesting that glutamate reuptake is necessary for the oscillatory release. GABAergic antagonists bicuculline, SR 95531, and picrotoxin increased the amplitude of the initial peak while they inhibit the frequency of oscillations. TTX had a similar effect. Strychnine, the blocker of glycine receptors did not affect the initial peak but strongly decreased the oscillations frequency. These inhibitory inputs onto the bipolar axon terminals shape and synchronize the oscillatory component.

Dendritic spread and functional coverage of starburst amacrine cells

The network of starburst amacrine cells plays a fundamental role in the neural circuitry underlying directional selectivity within the retina. Individual sectors of the starburst dendritic field are directionally selective by virtue of a mutually inhibitory relationship between starburst amacrine cells with overlapping dendrites. These features of the starburst amacrine cell network suggest that starburst cells regulate their dendritic overlap to ensure a uniform coverage of the retinal surface. The present study has compared the dendritic morphology of starburst amacrine cells in two different strains of mice that differ in starburst amacrine cell number. The A/J (A) strain contains about one-quarter fewer starburst amacrine cells than does the C57BL/6J (B6) strain, although the mosaics of starburst amacrine cells in both strains are comparably patterned. Dendritic field size, however, does not compensate for the difference in density, the A strain having a slightly smaller dendritic field relative to the B6 strain, yielding a significantly larger dendritic coverage factor for individual cells in the B6 strain. The area of the distal (output) annulus of the dendritic field occupies a comparable proportion of the overall field area in the two strains, but overlapping annuli establish a finer meshwork of co-fasciculating processes in the B6 strain. These results would suggest that the architecture of the dendritic network, rather than the overall size of the dendritic field, is dependent on the density of starburst amacrine cells.

Cellular Mechanisms for Direction Selectivity in the Retina

Direction selectivity represents a fundamental computation found across multiple sensory systems. In the mammalian visual system, direction selectivity appears first in the retina, where excitatory and inhibitory interneurons release neurotransmitter most rapidly during movement in a preferred direction. Two parallel sets of interneuron signals are integrated by a direction-selective ganglion cell, which creates a direction preference for both bright and dark moving objects. Direction selectivity of synaptic input becomes amplified by action potentials in the ganglion cell dendrites. Recent work has elucidated direction-selective mechanisms in inhibitory circuitry, but mechanisms in excitatory circuitry remain unexplained.

Displaced GAD65 amacrine cells of the guinea pig retina are morphologically diverse

The ganglion cell layer of mammalian retina contains numerous amacrine cells. Many belong to one type, the cholinergic starburst cell, but the other types have not been systematically identified. Using a new method to target sparsely represented cell types, we filled about 200 amacrine neurons in the ganglion cell layer of the guinea pig visual streak and identified 11 types. Ten of these resemble types identified in other species with somas in the inner nuclear layer, but one type has not been previously reported. Most of the types and nearly all the injected cells (95%) arborized low in the synaptic layer where they would co-stratify with various classes of ON ganglion cell. The displaced somas (7% of all amacrine cells) thus represent a heterogeneous pool, which are relatively accessible for study of their interactions with ON ganglion cells.

The Synaptic Mechanism of Direction Selectivity in Distal Processes of Starburst Amacrine Cells

Patch-clamp recordings revealed that distal processes of starburst amacrine cells (SACs) received largely excitatory synaptic input from the receptive field center and nearly purely inhibitory inputs from the surround during both stationary and moving light stimulations. The direct surround inhibition was mediated mainly by reciprocal GABA(A) synapses between opposing SACs, which provided leading and prolonged inhibition during centripetal stimulus motion. Simultaneous Ca(2+) imaging and current-clamp recording during apparent-motion stimulation further demonstrated the contributions of both centrifugal excitation and GABA(A/C)-receptor-mediated centripetal inhibition to the direction-selective Ca(2+) responses in SAC distal processes. Thus, by placing GABA release sites in electrotonically semi-isolated distal processes and endowing these sites with reciprocal GABA(A) synapses, SACs use a radial-symmetric center-surround receptive field structure to build a polar-asymmetric circuitry. This circuitry may integrate at least three levels of interactions--center excitation, surround inhibition, and reciprocal inhibitions that amplify the center--surround antagonism-to generate robust direction selectivity in the distal processes.

Dissociated GABAergic retinal interneurons exhibit spontaneous increases in intracellular calcium

Early in development, before the retina is responsive to light, neurons exhibit spontaneous activity. Recently it was demonstrated that starburst amacrine cells, a unique class of neurons that secretes both GABA and acetylcholine, spontaneously depolarize. Networks comprised of spontaneously active starburst cells initiate correlated bursts of action potentials that propagate across the developing retina with a periodicity on the order minutes. To determine whether other retinal interneurons have similar "pacemaking" properties, we have utilized cultures of dissociated neurons from the rat retina. In the presence of antagonists for fast neurotransmitter receptors, distinct populations of neurons exhibited spontaneous, uncorrelated increases in intracellular calcium concentration. These increases in intracellular calcium concentration were sensitive to tetrodotoxin, indicating they are mediated by spontaneous membrane depolarizations. By combining immunofluorescence and calcium imaging, we found that 44% of spontaneously active neurons were GABAergic and included starburst amacrine cells. Whole cell voltage clamp recordings in the absence of antagonists for fast neurotransmitters revealed that after 7 days in culture, individual retinal neurons receive bursts of GABA-A receptor mediated synaptic input with a periodicity similar to that measured in spontaneously active GABAergic neurons. Low concentrations of GABA-A receptor antagonists did not alter the inter-burst interval despite significant reduction of post-synaptic current amplitude, indicating that pacemaker activity of GABAergic neurons was not influenced by network interactions. Together, these findings indicate that spiking GABAergic interneurons can function as pacemakers in the developing retina.

Modeling Starburst Cells’ GABA B Receptors and Their Putative Role in Motion Sensitivity

Neal and Cunningham (Neal, M. J., and J. R. Cunningham. 1995. J. Physiol. (Lond.). 482:363–372) showed that GABA B agonists and glycinergic antagonists enhance the light-evoked release of retinal acetylcholine. They proposed that glycinergic cells inhibit the cholinergic Starburst amacrine cells and are in turn inhibited by GABA through GABA B receptors. However, as recently shown, glycinergic cells do not appear to have GABA B receptors. In contrast, the Starburst amacrine cell has GABA B receptors in a subpopulation of its varicosities. We thus propose an alternate model in which GABA B-receptor activation reduces the release of ACh from some dendritic compartments onto a glycinergic cell, which then feeds back and inhibits the Starburst cell. In this model, the GABA necessary to make these receptors active comes from the Starburst cell itself, making them autoreceptors. Computer simulations of this model show that it accounts quantitatively for the Neal and Cunningham data. We also argue that GABA B receptors could work to increase the sensitivity to motion over other stimuli.

Symmetric Interactions Within a Homogeneous Starburst Cell Network Can Lead to Robust Asymmetries in Dendrites of Starburst Amacrine Cells

Starburst amacrine cells in the mammalian retina respond asymmetrically to movement along their dendrites; centrifugal movement elicits stronger responses in each dendrite than centripetal movement. It has been suggested that the asymmetrical response can be attributed to intrinsic properties of the processes themselves. But starburst cells are known to release and have receptors for both GABA and acetylcholine. We tested whether interactions within the starburst cell network can contribute to their directional response properties. In a computational model of interacting starburst amacrine cells, we simulated the response of individual dendrites to moving light stimuli. By setting the model parameters for "synaptic connection strength" (cs) to positive or negative values, overlapping starburst dendrites could either excite or inhibit each other. For some values of cs, we observed a very robust inward/outward asymmetry of the starburst dendrites consistent with the reported physiological findings. This is the case, for example, if a starburst cell receives inhibition from other starburst cells located in its surround. For other values of cs, individual dendrites can respond best either to inward movement or respond symmetrically. A properly wired network of starburst cells can therefore account for the experimentally observed asymmetry of their response to movement, independent of any internal biophysical or biochemical properties of starburst cell dendrites.

A transient network of intrinsically bursting starburst cells underlies the generation of retinal waves

Pharmacologically isolated starburst amacrine cells (SACs) in perinatal rabbit retinas spontaneously generated semiperiodic calcium spikes and long-lasting after-hyperpolarizations (AHPs), mediated by calcium-activated, cyclic AMP-sensitive potassium currents. These AHPs, rather than a depletion of neurotransmitters (as was previously believed), produced the refractory period of spontaneous retinal waves and set the upper limit of the wave frequency. Each SAC received inputs from roughly 10-30 neighboring SACs during a wave. These inputs synchronized and reshaped the intrinsic bursts to produce network oscillations at a rhythm different from that of individual SACs. With maturation, the semiperiodic bursts in SACs disappeared, owing to reduced intrinsic excitability and increased network inhibition. Thus, retinal waves are generated by a transient and specific network of cell-autonomous oscillators synchronized by reciprocally excitatory connections.

Spontaneous Oscillatory Activity of Starburst Amacrine Cells in the Mouse Retina

Using patch-clamp techniques, we investigated the characteristics of the spontaneous oscillatory activity displayed by starburst amacrine cells in the mouse retina. At a holding potential of -70 mV, oscillations appeared as spontaneous, rhythmic inward currents with a frequency of approximately 3.5 Hz and an average maximal amplitude of approximately 120 pA. Application of TEA, a potassium channel blocker, increased the amplitude of oscillatory currents by >70% but reduced their frequency by approximately 17%. The TEA effects did not appear to result from direct actions on starburst cells, but rather a modulation of their synaptic inputs. Oscillatory currents were inhibited by 6-cyano-7-nitroquinoxalene-2,3-dione (CNQX), an antagonist of AMPA/kainate receptors, indicating that they were dependent on a periodic glutamatergic input likely from presynaptic bipolar cells. The oscillations were also inhibited by the calcium channel blockers cadmium and nifedipine, suggesting that the glutamate release was calcium dependent. Application of AP4, an agonist of mGluR6 receptors on on-center bipolar cells, blocked the oscillatory currents in starburst cells. However, application of TEA overcame the AP4 blockade, suggesting that the periodic glutamate release from bipolar cells is intrinsic to the inner plexiform layer in that, under experimental conditions, it can occur independent of photoreceptor input. The GABA receptor antagonists picrotoxin and bicuculline enhanced the amplitude of oscillations in starburst cells prestimulated with TEA. Our results suggest that this enhancement was due to a reduction of a GABAergic feedback inhibition from amacrine cells to bipolar cells and the resultant increased glutamate release. Finally, we found that some ganglion cells and other types of amacrine cell also displayed rhythmic activity, suggesting that oscillatory behavior is expressed by a number of inner retinal neurons.

The many roles of starburst amacrine cells

Starburst amacrine cells release two classical neurotransmitters, ACh and GABA. In a tour de force of paired-cell recording, Zheng et al. now show that the starburst cells are mutually excitatory during early development but mutually inhibitory in adult animals. The change occurs by remodeling of both the cholinergic and the GABAergic synapses between starburst cells. The finding gives a precise mechanistic basis for the developmental waves of activity in the retina.

Directional Selectivity Is Formed at Multiple Levels by Laterally Offset Inhibition in the Rabbit Retina

The excitatory and inhibitory inputs to directionally selective (DS) ganglion cells are themselves directionally selective. Directionality is achieved because excitation is reduced during null-direction movement along a GABAergic pathway. Inhibition is reduced during preferred-direction movement along a pathway that includes cholinergic synapses. Both excitation and inhibition are made directional by laterally offset inhibitory signals similar to the spatial offset of the direct inhibitory input to the DS cell dendrites. Thus, spatially offset lateral inhibition generates directionality at three different levels in the DS circuitry. We also found that for stimuli falling within the dendritic field, cholinergic input is delivered to the OFF but not the ON dendrites. Cholinergic pathways from outside the dendritic field reach both ON and OFF dendrites, but both of these pathways are normally inactivated by GABAergic synapses.

Starburst Network Goes from Excitatory to Inhibitory

During development, the action of γ-aminobutyric acid (GABA) changes from an excitatory neurotransmitter to an inhibitory one. Zheng et al. show that, in starburst neurons of the developing mammalian retina [rabbits, embryonic day 29 (E29) to postnatal day 1 (P1)], a single neuron releases both acetylcholine (ACh) and GABA. The nicotinic ACh synaptic activity diminished during development so that after P11, the time when the rabbits' eyes opened, only GABAergic responses were detectable. At the same time that these nicotinic cholinergic synapses were being eliminated, the starburst cell morphology was also changing to one with numerous varicosities. Pharmacological analysis coupled with electrophysiological recordings indicated that the spontaneous retinal waves associated with the starburst cells at the early stages of development (E30) were the result of mutual nicotinic cholinergic excitation among the starburst cells. In cells from P8 animals, GABA input became inhibitory, and the retinal waves were insensitive to nicotinic antagonists and were then mediated by glutamatergic synaptic interactions. After eye opening, GABA mediated light-evoked synaptic inhibition among neighboring starburst cells. Thus, the conversion of GABA synaptic activity from excitatory to stimulatory and the elimination of nicotinic ACh activity change the character of the retinal starburst synaptic network from one with recurrent spontaneous rhythmic activity to a stable network. J.-J. Zheng, S. Lee, Z. J. Zhou, A developmental switch in the excitability and function of the starburst network in the mammalian retina. Neuron 44 , 851-864 (2004). [Online Journal]