Biomedical subjects

Stewart A Bloomfield

Publications and source records attributed to Stewart A Bloomfield.

10 recordsLinked to original sources

PubMed2006

Light-induced changes in spike synchronization between coupled ON direction selective ganglion cells in the mammalian retina.

Although electrical coupling via gap junctions is prevalent among ganglion cells in the vertebrate retina, there have been few direct studies of their influence on the light-evoked signaling of these cells. Here, we describe the pattern and function of coupling between the ON direction selective (DS) ganglion cells, a unique subtype whose signals are transmitted to the accessory optic system (AOS) where they initiate the optokinetic response. ON DS cells are coupled indirectly via gap junctions made with a subtype of polyaxonal amacrine cell. This coupling underlies synchronization of the spontaneous and light-evoked spike activity of neighboring ON DS cells. However, we find that ON DS cell pairs show robust synchrony for all directions of stimulus movement, except for the null direction. Null stimulus movement evokes a GABAergic inhibition that temporally shifts firing of ON DS cell neighbors, resulting in a desynchronization of spike activity. Thus, detection of null stimulus movement appears key to the direction selectivity of ON DS cells, evoking both an attenuation of spike frequency and a desynchronization of neighbors. We posit that active desynchronization reduces summation of synaptic potentials at target AOS cells and thus provides a secondary mechanism by which ON DS cell ensembles can signal direction of stimulus motion to the brain.

Action Potentials↗

PubMed2005

Morphology and tracer coupling pattern of alpha ganglion cells in the mouse retina.

Alpha cells are a type of ganglion cell whose morphology appears to be conserved across a number of mammalian retinas. In particular, alpha cells display the largest somata and dendritic arbors at a given eccentricity and tile the retina as independent on- (ON) and off-center (OFF) subtypes. Mammalian alpha cells also express a variable tracer coupling pattern, which often includes homologous (same cell type) coupling to a few neighboring alpha cells and extensive heterologous (different cell type) coupling to two to three amacrine cell types. Here, we use the gap junction-permeant tracer Neurobiotin to determine the architecture and coupling pattern of alpha cells in the mouse retina. We find that alpha cells show the same somatic and dendritic architecture described previously in the mammal. However, alpha cells show varied tracer coupling patterns related to their ON and OFF physiologies. ON alpha cells show no evidence of homologous tracer coupling but are coupled heterologously to at least two types of amacrine cell whose somata lie within the ganglion cell layer. In contrast, OFF alpha cells are coupled to one another in circumscribed arrays as well as to two to three types of amacrine cell with somata occupying the inner nuclear layer. We find that homologous coupling between OFF alpha cells is unaltered in the connexin36 (Cx36) knockout (KO) mouse retina, indicating that it is not dependent on Cx36. However, a subset of the heterologous coupling of ON alpha cells and all the heterologous coupling of OFF alpha cells are eliminated in the KO retina, suggesting that Cx36 comprises most of the junctions made with amacrine cells.

Amacrine Cells↗

PubMed2004

Convergence and segregation of the multiple rod pathways in mammalian retina.

Using a multidisciplinary approach, we demonstrate that three different pathways are responsible for the transmission of rod signals across the mouse retina. Each pathway serves a primarily nonoverlapping range of stimulus intensities, with ganglion cells receiving either segregated or convergent inputs. For both on-center (ON) and off-center (OFF) ganglion cells, the primary rod pathway carries signals with the lowest threshold, whereas the secondary rod pathway is less sensitive by approximately 1 log unit. In addition, OFF signaling uses a tertiary rod pathway that is approximately 1 log unit less sensitive than the secondary. Although some ganglion cells received rod inputs exclusively from one of the pathways, others showed convergent inputs. Using pharmacological and genetic approaches, we defined classes of ON and OFF ganglion cells for which the scotopic inputs derive only from the primary pathway or from both primary and secondary pathways. In addition, we observed a class of OFF ganglion cell receiving mixed input from primary and tertiary pathways. Interestingly, OFF ganglion cells receiving convergent inputs from all three rod pathways or from the secondary and tertiary pathways together were never observed. Overall, our data show a complex arrangement of convergence and segregation of rod inputs to ganglion cells in the mammalian retina.

Action Potentials↗

PubMed2004

A unique role for Kv3 voltage-gated potassium channels in starburst amacrine cell signaling in mouse retina.

Direction-selective retinal ganglion cells show an increased activity evoked by light stimuli moving in the preferred direction. This selectivity is governed by direction-selective inhibition from starburst amacrine cells occurring during stimulus movement in the opposite or null direction. To understand the intrinsic membrane properties of starburst cells responsible for direction-selective GABA release, we performed whole-cell recordings from starburst cells in mouse retina. Voltage-clamp recordings revealed prominent voltage-dependent K(+) currents. The currents were mostly blocked by 1 mm TEA, activated rapidly at voltages more positive than -20 mV, and deactivated quickly, properties reminiscent of the currents carried by the Kv3 subfamily of K+ channels. Immunoblots confirmed the presence of Kv3.1 and Kv3.2 proteins in retina and immunohistochemistry revealed their expression in starburst cell somata and dendrites. The Kv3-like current in starburst cells was absent in Kv3.1-Kv3.2 knock-out mice. Current-clamp recordings showed that the fast activation of the Kv3 channels provides a voltage-dependent shunt that limits depolarization of the soma to potentials more positive than -20 mV. This provides a mechanism likely to contribute to the electrical isolation of individual starburst cell dendrites, a property thought essential for direction selectivity. This function of Kv3 channels differs from that in other neurons where they facilitate high-frequency repetitive firing. Moreover, we found a gradient in the intensity of Kv3.1b immunolabeling favoring proximal regions of starburst cells. We hypothesize that this Kv3 channel gradient contributes to the preference for centrifugal signal flow in dendrites underlying direction-selective GABA release from starburst amacrine cells

Amacrine Cells↗

PubMed2004

Control of late off-center cone bipolar cell differentiation and visual signaling by the homeobox gene Vsx1.

Retinal bipolar cells are interneurons that transmit visual signals from photoreceptors to ganglion cells. Although the visual pathways mediated by bipolar cells have been well characterized, the genes that regulate their development and function are largely unknown. To determine the role in bipolar cell development of the homeobox gene Vsx1, whose retinal expression is restricted to a major subset of differentiating and mature cone bipolar (CB) cells, we targeted the gene in mice. Bipolar cell fate was not altered in the absence of Vsx1 function, because the pan-bipolar markers Chx10 and Ret-B1 continued to be expressed in inner nuclear layer neurons labeled by the Vsx1-targeting reporter gene, tauLacZ. The specification, number, and gross morphology of the subset of on-center and off-center (OFF)-CB cells defined by tauLacZ expression from the Vsx1 locus were also normal in Vsx1(tauLacZ)/Vsx1(tauLacZ) mice. However, the terminal differentiation of OFF-CB cells in the retina of Vsx1(tauLacZ)/Vsx1(tauLacZ) mice was incomplete, as demonstrated by a substantial reduction in the expression of at least four markers (recoverin, NK3R, Neto1, and CaB5) for these interneurons. These molecular abnormalities were associated with defects in retinal function and documented by electroretinography and in vitro ganglion cell recordings specific to cone visual signaling. In particular, there was a general reduction in the light-mediated activity of OFF, but not on-center, ganglion cells. Thus, Vsx1 is required for the late differentiation and function of OFF-CB cells and is associated with a heritable OFF visual pathway-specific retinal defect.

Animals↗

PubMed2004

Function and plasticity of homologous coupling between AII amacrine cells.

The AII amacrine cells are critical elements in the primary rod pathway of the mammalian retina, acting as an obligatory conduit of rod signals to both on- and off-center ganglion cells. In addition to the chemical synaptic circuitry they subserve, AII cells form two types of electrical synapses corresponding to gap junctions formed between neighboring AII cells as well as junctions formed between AII cells and on-center cone bipolar cells. Our recent results indicate that coupling between AII cells and cone bipolar cells forms an obligatory synapse for transmission of scotopic visual signals to on-center ganglion cells. In contrast, AII-AII cell coupling acts to maintain the sensitivity of the primary rod pathway by allowing for summation of synchronous activity and the attenuation of asynchronous background noise. Further, the conductance of AII-AII cell gap junctions is highly dynamic, regulated by ambient light conditions, thereby preserving the fidelity of rod signaling over the scotopic operating range from starlight to twilight.

Amacrine Cells↗

PubMed2003

Gap junctional coupling underlies the short-latency spike synchrony of retinal alpha ganglion cells.

We examined whether coupling between neighboringalpha-type ganglion cells (alpha-GCs) in the rabbit retina underlies their synchronous spike activity. Simultaneous recordings were made from arrays of alpha-GCs to determine the synchrony of both spontaneous and light-evoked spike activity. One cell within each array was then injected with the biotinylated tracer Neurobiotin to determine which of the cells were coupled via gap junctions. Cross-correlation analyses indicated that neighboring off-center alpha-GCs maintain short-latency (approximately 2.5 msec) synchronous spiking, whereas the spontaneous spike activities of on-centeralpha-GC neighbors are not correlated. Without exception, those off-centeralpha-GCs showing synchronous spiking were found to be tracer coupled to both amacrine cells and neighboring off-centeralpha-GCs. In contrast, on-center alpha-GCs were never tracer coupled. Furthermore, whereas spikes initiated in an off-center alpha-GC with extrinsic current injection resulted in short-latency synchronized spiking in neighboring off-center alpha-GCs, this was never seen between on-center alpha-GCs. These results indicate that electrical coupling via gap junctions underlies the short-latency concerted spike activity of neighboring alpha-GCs.

Action Potentials↗

PubMed2002

Axonal neurofilament-H immunolabeling in the rabbit retina.

A monoclonal antibody directed at the multiphosphorylated epitope of axonal neurofilament-H (NF-H) was used to label axon-like fibers in the rabbit retina. NF-H-immunopositive fibers were found in the outer plexiform layer (OPL), inner plexiform layer (IPL), and optic fiber layer (OFL). The morphological characteristics of the labeled processes identified those in the OPL as horizontal cell axons and axon terminals and fibers in the OFL as axons of ganglion cells. The NF-H-positive profiles in the OPL formed a subset of horizontal cell processes labeled for calbindin. In the IPL, NF-H-immunoreactive profiles lay at all levels but were detected most often in the middle strata, 2-4. Occasionally, we observed NF-H-immuoreactive processes emerging from the IPL and entering either the GCL or the inner nuclear layer (INL). The labeled fibers in the IPL were typically very thin, less than 1 microm in diameter, and could often be followed for over 1 mm as they ran laterally across the retina. Cell bodies were never labeled by the immunoserum. To identify the NF-H-immunopositive fibers in the IPL, standard immunocytochemical double-labeling techniques were applied, using antibodies directed against several neurotransmitters or modulators thought to be expressed by axon-bearing amacrine cells. The NF-H-positive processes in the IPL were found to correspond to those labeled for tyrosine hydroxylase, somatostatin, substance P, and NADPH diaphorase activity. However, the NF-H labels did not colocalize with those against the vasoactive intestinal peptide-associated protein PHM27. Our results indicate that putative axons in the retina possess the multiphosphorylated NF-H protein found within classic axons in the central nervous system. These results thus support the idea that certain subtypes of amacrine and horizontal cells maintain true axons in the mammalian retina.

Amacrine Cells↗

PubMed2002

Connexin36 is essential for transmission of rod-mediated visual signals in the mammalian retina.

To examine the functions of electrical synapses in the transmission of signals from rod photoreceptors to ganglion cells, we generated connexin36 knockout mice. Reporter expression indicated that connexin36 was present in multiple retinal neurons including rod photoreceptors, cone bipolar cells, and AII amacrine cells. Disruption of electrical synapses between adjacent AIIs and between AIIs and ON cone bipolars was demonstrated by intracellular injection of Neurobiotin. In addition, extracellular recording in the knockout revealed the complete elimination of rod-mediated, on-center responses at the ganglion cell level. These data represent direct proof that electrical synapses are critical for the propagation of rod signals across the mammalian retina, and they demonstrate the existence of multiple rod pathways, each of which is dependent on electrical synapses.

Alkaline Phosphatase↗

PubMed2002

Feedback inhibition in the inner plexiform layer underlies the surround-mediated responses of AII amacrine cells in the mammalian retina.

Intracellular recordings were made from narrow-field, bistratified AII amacrine cells in the isolated, superfused retina-eyecup of the rabbit. Pharmacological agents were applied to neurons to dissect the synaptic pathways subserving AII cells so as to determine the circuitry generating their off-surround responses. Application of the GABA antagonists, picrotoxin, bicuculline and 1,2,5,6-tetrahydropyridine-4-yl methylphosphinic acid (TPMPA) all increased the on-centre responses of AII amacrine cells, but attenuated the off-surround activity. At equal concentrations, picrotoxin was approximately twice as effective as bicuculline or TPMPA in modifying the response activity of AII amacrine cells. These results indicate that the mechanism underlying surround inhibition of AII amacrine cells includes activation of both GABA(A) and GABA(C) receptors in an approximately equal ratio. Application of the GABA antagonists also increased the size of on-centre receptive fields of AII amacrine cells. Again, picrotoxin was most effective, producing, on average, a 54 % increase in the size of the receptive field, whereas bicuculline and TPMPA produced comparable 34 and 33 % increases, respectfully. Application of the voltage-gated sodium channel blocker TTX produced effects on AII amacrine cells qualitatively similar to those of the GABA blockers. Intracellular application of the chloride channel blocker 4,4'-dinitro-stilbene-2,2'-disulphonic acid (DNDS) abolished the direct effects of GABA on AII amacrine cells. Moreover, DNDS increased the amplitude of both the on-centre and off-surround responses. The failure of DNDS to block the off-surround activity indicates that it is not mediated by direct GABAergic inhibition. Taken together, our results suggest that surround receptive fields of AII amacrine cells are generated indirectly by the GABAergic, reciprocal feedback synapses from S1/S2 amacrine cells to the axon terminals of rod bipolar cells.

Animals↗