Synaptic transmission between photoreceptors and horizontal cells in the turtle retina.

Low calcium, high magnesium, and cobalt hyperpolarize the horizontal cell membrane and suppress the response to light, but only partially affect the response of receptor cells. These observations are consistent with the interpretation that a depolarizing transmitter is released by photoreceptors in darkness. The hyperpolarizing response to light of the horizontal cells would then result from a reduction in the amount of transmitter released.

Variations of the visual responses of the superior colliculus in relation to body roll.

A large percentage of the directional units of the superior colliculus of the curarized cat modify their response to a particular moving visual stimulus as a function of the position of rotation of the animal about its longitudinal axis.

The bicentennial of the Voltaic battery (1800-2000): the artificial electric organ.

Alessandro Volta invented the electric battery at the end of 1799 and communicated his invention to the Royal Society of London in 1800. The studies that led him to develop this revolutionary device began in 1792, after Volta read the work of Luigi Galvani on the existence of an intrinsic electricity in living organisms. During these studies, Volta obtained a series of results of great physiological relevance, which led him to anticipate some important ideas that marked the inception of modern neuroscience. These results have been obscured by a cultural tradition that has seen Volta exclusively as a physicist, lacking interest for biological problems and opposed in an irreversible way to the physiologist, Luigi Galvani.

Luigi Galvani and animal electricity: two centuries after the foundation of electrophysiology.

Luigi Galvani and his famous experiments on frogs carried out in the second half of the 18th century belong more to legend than to the history of science. Galvani not only laid the foundations of a new science, electrophysiology, but also opened the way for the invention of the electric battery, and thus for the development of the physical investigations of electricity. However, in spite of the widespread celebration of his work, Galvani's scientific endeavours have been largely misrepresented in the history of science. The scholar of Bologna has a stereotyped image as an 'occasional' scientist, who started his studies by chance, largely ignored the scientific theories of his time and wandered aimlessly in mental elaborations until the physicist of Pavia, Alessandro Volta, entered the field, correctly interpreted Galvani's results and eventually developed the electric battery. With the present understanding of electrical phenomena in excitable membranes, it is now time to reconsider the real matter raised by Galvani's discoveries and by his hypothesis of an intrinsic 'animal electricity', and to make a clearer evaluation of a revolutionary phase of scientific progress.

Calcium-independent synaptic transmission: artifact or fact?

The release of neurotransmitters at classical chemical synapses occurs via Ca2+ influx through voltage-dependent Ca2+ channels, which are opened following depolarization of presynaptic terminals. However, owing to a persistence or increase in the amount of transmitter released in preparations containing low concentrations of Ca2+, it has been proposed that transmitter release could also occur through a Ca(2+)-independent, carrier-mediated process. On the other hand, lowering extracellular [Ca2+] can actually promote Ca2+ influx through voltage-activated Ca2+ channels via a modification of the surface potential of plasma membranes. Therefore, the proposed Ca(2+)-independent transmitter release could be re-accommodated within the framework of the Ca2+ hypothesis of synaptic transmission by taking into account the surface-charge effects.

Calcium-independent release of neurotransmitter in the retina: a "copernican" viewpoint change.

The release of synaptic transmitter in chemical synapses is brought about by Ca2+ influx through voltage-dependent Ca2+ channels opened by depolarisation of presynaptic terminals. However, in some preparations transmitter release persists or increases in low-Ca2+ media, and it has therefore been proposed that transmitter release could also occur through a Ca2+-independent, carrier mediated process. In particular it has been suggested that this may be the case for synaptic transmission between photoreceptors and second order neurones of the vertebrate retina. From our recent experiments on synaptic transmission from photoreceptors to horizontal cells of turtle and salamander retinas, it appears that lowering extracellular Ca2+ can actually promote Ca2+ influx through voltage-activated Ca2+ channels via a modification of surface potential of plasma membranes. On the basis of this apparently paradoxical effect of low Ca2+ media, it is possible to reaccommodate the so-called Ca2+-independent release within the framework of Ca2+-dependent synaptic transmission without invoking unconventional mechanisms.

Involvement of D1 and D2 Dopamine Receptors in the Control of Horizontal Cell Electrical Coupling in the Turtle Retina.

We studied the actions of D1 and D2 dopamine agonists and antagonists on the coupling of horizontal cell axons in the turtle retina by a combination of pharmacological and electrophysiological methods. Both D1 and D2 receptors were identified in membrane fractions by radioligand binding using [3H]-SCH 23390 and [3H]-spiperone, respectively. The KD of both receptor classes were identical (0.21 nM) but D1 receptor density exceeded that of D2 receptors by more than four-fold. D1 agonists increased the activity of adenylate cyclase in a dose-dependent manner, whereas D2 agonists were without significant effect by themselves, nor did D2 antagonists block the D1-mediated increase in adenylate cyclase activity. Intracellular recordings and Lucifer Yellow dye injections were used to characterize the modifications of the receptive field profile of horizontal cell axons (H1AT) exposed to different pharmacological agents. Dopamine or D1 agonists (0.05 - 10 microM) induced a marked constriction of the H1AT receptive field, whereas D2 agonists elicited a small expansion of the receptive field. However, in the presence of a D1 antagonist, as well as IBMX to inhibit phosphodiesterase, D2 agonists (10 - 70 microM) induced a marked increase in the receptive field profile. These results indicate that both D1 and D2 dopamine receptors play a role in shaping the receptive field profile of the horizontal cell axon terminal in the turtle retina.

Pre- and post-synaptic effects of manipulating surface charge with divalent cations at the photoreceptor synapse.

Persistence of horizontal cell (HC) light responses in extracellular solutions containing low Ca2+ plus divalent cations to block Ca2+ currents (ICa) has been attributed to Ca2+-independent neurotransmission. Using a retinal slice preparation to record both ICa and light responses, we demonstrate that persistence of HC responses in low [Ca2+]o can instead be explained by a paradoxical increase of Ca2+ influx into photoreceptor terminals arising from surface charge-mediated shifts in ICa activation. Consistent with this explanation, application of Zn2+ or Ni2+ caused a hyperpolarizing block of HC light responses that was relieved by lowering [Ca2+]o. The same concentrations of Zn2+ and Ni2+ reduced the amplitude of ICa at the rod dark potential and this reduction was relieved by a hyperpolarizing shift in voltage dependence induced by lowering [Ca2+]o. Block of ICa by Mg2+, which has weak surface charge effects, was not relieved by low [Ca2+]o. Recovery of HC responses in low [Ca2+]o was assisted by enhancement of rod light responses. To bypass light stimulation, OFF bipolar cells were stimulated by steps to -40 mV applied to presynaptic rods during simultaneous paired recordings. Consistent with surface charge theory, the post-synaptic current was inhibited by Zn2+ and this inhibition was relieved by lowering [Ca2+]o. Nominally divalent-free media produced inversion of HC light responses even though rod light responses remained hyperpolarizing; HC response inversion can be explained by surface charge-mediated shifts in ICa. In summary, HC light responses modifications induced by low divalent cation solutions can be explained by effects on photoreceptor light responses and membrane surface charge without necessitating Ca2+-independent neurotransmission. Furthermore, these results suggest that surface charge effects accompanying physiological changing divalent cation levels in the synaptic cleft may provide a means for modulating synaptic output from photoreceptors.

Autoradiographic distribution of peripheral benzodiazepine receptors in the retina of the albino rabbit, Lepus cunicula.

The distribution of peripheral benzodiazepine receptors (PBRs) in the retina of the albino rabbit, Lepus cunicula, was studied by autoradiography using [3H]-PK11195, a isoquinoline carboxamide, as a tracer. Autoradiograms obtained by directly placing the slides containing the retina sections on tritium-sensitive film provide evidence for the presence of PBRs in rabbit retina. Furthermore, the dark field examination of photomicrographs taken from autoradiograms showed two dense horizontal bands corresponding to the outer and inner photoreceptor segments, and to the inner plexiform layer. The retinal regions where [3H]-PK11195 binding was more dense are rich in mitochondria, suggesting that as in other neuronal tissues, retinal PBRs are involved in the mitochondrial activity.

Animal electricity and the birth of electrophysiology: the legacy of Luigi Galvani.

Preceded by a companion paper on Galvani's life, this article is written on the occasion of the bicentenary of the death of Luigi Galvani. From his studies on the effects of electricity on frogs, the scientist of Bologna derived the hypothesis that animal tissues are endowed with an intrinsic electricity that is involved in fundamental physiological processes such as nerve conduction and muscle contraction. Galvani's work swept away from life sciences mysterious fluids and elusive entities like "animal spirits" and led to the foundation of a new science, electrophysiology. Two centuries of research work have demonstrated how insightful was Galvani's conception of animal electricity. Nevertheless, the scholar of Bologna is still largely misrepresented in the history of science, because the importance of his researches seems to be limited to the fact that they opened the paths to the studies of the physicist Alessandro Volta, which culminated in 1800 with the invention of the electric battery. Volta strongly opposed Galvani's theories on animal electricity. The matter of the scientific controversy between Galvani and Volta is examined here in the light of two centuries of electrophysiological studies leading to the modern understanding of electrical excitability in nerve and muscle. By surveying the work of scientists such as Nobili, Matteucci, du Bois-Reymond, von Helmholtz, Bernstein, Hermann, Lucas, Adrian, Hodgkin, Huxley, and Katz, the real matter of the debate raised by Galvani's discoveries is here reconsidered. In addition, a revolutionary phase of the 18th century science that opened the way for the development of modern neurosciences is reevaluated.

Marcello Malpighi and the difficult birth of modern life sciences.

All his life, Marcello Malpighi (1628-1694), the founder of modern microscopic anatomy, was unwillingly involved in difficult debates within a reactionary medical milieu that questioned the significance of modern science and its utility to medicine. Malpighi's responses to his detractors, included in posthumous works first published in 1697 by the Royal Society, offer an important insight into a critical phase of scientific progress in the 17th century and help to reveal the prevailing conception of science. In some ways, Malpighi's views predate important ideas in modern biology.

Processing of visual signals in vertebrate photoreceptors.

Photoreceptors of the vertebrate retina hyperpolarize in response to illumination. The conductance changes in the plasma membrane associated with the electrical response are the final step of chain of events initiated by light absorption at the outer segment of the visual cell. The mechanism whereby the free energy of photons in converted into neural information is largely unknown. Present knowledge is consistent with the idea that an internal transmitter is modulated by light and modifies the ionic permeability of the plasma membrane. As to the identity of the internal messenger two candidates have been proposed: Ca2+ and cyclic GMP respectively. Increasing evidence suggests that both substances may be involved in the process of phototransduction. The electrical response of photoreceptors does not simply reflect the light absorbed by the cell: complex interactions occurring between adjacent photoreceptors and between photoreceptors and second order neurons cooperate with the initial process in determining the final shape of the receptor message. Recurrent interactions involve particularly cones: their membrane potential can be modified at least by three distinct mechanisms; i) by light absorption at their outer segment; ii) by light absorption at the outer segment of neighbouring cones, and iii) by potential changes occurring in horizontal cells.

Retinal horizontal cells: old cells, old experiments, new results.

The study of neural interactions in the vertebrate retina carried out after the pioneering studies of Svaetichin has provided important information on the functioning of nerve circuits in the central nervous system. Recently we have investigated the effects of changes of divalent cation concentration on the synaptic transmission between cones and horizontal cells of the turtle retina. Our results seemed apparently in contrast with the classical Ca2(+)-hypothesis of chemical synaptic transmission. Application of low Ca2+ media resulted in a recovery of synaptic transmission after application of divalent cations such as Ca2+, Zn2+ and Ni2+ traditionally considered as Ca2+ channel antagonists. Moreover, in the absence of exogenous divalent cations, low Ca2+ could result in an increase of transmitter release particularly if Mg2+ was omitted from the perfusing medium. These apparently paradoxical results can be reconciled with the postulates of the Ca2(+)-hypothesis of synaptic transmission by taking into account the effects of divalent cations on the fixed charges present at the external surface of cell membrane. It is possible that a similar interpretation could also account for the so-called "Ca2(+)-independent" transmission in other structures of the nervous system.

Biological machines: from mills to molecules.

Although scientific progress is usually represented as being linear, it may, in fact, have a cyclical character--some discoveries may be forgotten or lost (at least temporarily), and themes may reappear through the centuries. Consider, for example, the concept of 'molecular machines', from the exciting phase of research that flourished in the seventeenth century, to the idea of machines that is at centre stage in modern cell biology.

Center-surround antagonistic organization in small-field luminosity horizontal cells of turtle retina.

1. The spatial properties of the small-field luminosity horizontal cell (L2-HC) were investigated in the retina of the turtle by using circular or annular light stimuli of varying dimensions. 2. The amplitude of the hyperpolarizing response induced by dim light spots decreased when the diameter of the spot was increased beyond about 1,000 micrometer. 3. With bright-light stimuli the peak amplitude of the light responses increased monotonically when increasing the illuminated area, but a delayed antagonistic effect appeared with spots of more than 1,000-micrometer diameter. 4. Depolarizing responses were observed if the periphery of the receptive field was stimulated with annuli of light in the presence of central background illumination. 5. The interaction of the inputs converging on the L2-HC from the peripheral and central regions of the receptive field was found to be nonlinear. The same peripheral stimulation could result either in an enhancement or in a depression of the central response according to the intensity of the central illumination. 6. By comparing the receptive-field properties of the L2-HC and the red cones a model is proposed that explains the antagonistic surround mechanism of the L2-HC on the basis of the antagonistic peripheral mechanism in cones. '

Morphological changes induced in turtle retinal neurons by exposure to 6-hydroxydopamine and 5,6-dihydroxytryptamine.

Following intraocular injection of the dopamine neurotoxin 6-hydroxydopamine (10-50 micrograms on two successive days in a Ringer vehicle containing ascorbate and pargyline) and an incubation period of 1 to 18 days, degeneration was noted in presumptive amacrine cells in the retina of the turtle, Pseudemys scripta elegans. Injection of vehicle alone produced no effect. Affected perikarya initially showed swollen mitochondria, lysosomes and distended cisternae. At later stages the cells took on a darkened appearance. In contrast, affected amacrine processes in the inner plexiform layer became markedly distended and lost their cytoplasmic contents, resulting in empty, very swollen profiles. No degeneration was noted distal to the affected cell bodies, i.e. the affected cells were not interplexiform neurons. Cells lesioned by 6-hydroxydopamine were shown to accumulate [3H]dopamine. Intraocular administration of 5,6-dihydroxytryptamine (a single dose of 10-40 micrograms in the same vehicle) followed by 4-6 days incubation resulted in a marked darkening of certain bipolar cell axon terminals, cell bodies and Landolt's clubs. The toxic effects of 5,6-dihydroxytryptamine were blocked by zimelidine, a serotonin uptake blocker. Thus, these two neurotoxins have different targets in the turtle retina. At the highest dose tested, however, 6-hydroxydopamine did produce degenerative changes in the presumed serotonergic bipolar cell.

Dopaminergic mechanisms underlying the reduction of electrical coupling between horizontal cells of the turtle retina induced by d-amphetamine, bicuculline, and veratridine.

Previous studies have shown that dopamine, bicuculline, or d-amphetamine reduce the electrical and dye-coupling between the axon terminals of the horizontal cells of the turtle retina (see Piccolino et al., 1984). In the present study we observed similar effects following the application of veratridine. The actions of all these drugs were prevented by dopamine antagonists acting on D1 receptors such as flupenthixol and SCH 23390. However, in contrast to dopamine, the actions of d-amphetamine, bicuculline, and veratridine were attenuated or abolished by pharmacological agents (such as 6-OH-dopamine, alpha-methyl-p-tyrosine, or reserpine) known to reduce the release of dopamine from dopaminergic neurons. Moreover, the actions of veratridine and bicuculline were prevented by tetrodotoxin, indicating that one or more neurons in the dopamine pathway are spike-generating. We conclude that d-amphetamine, bicuculline, and veratridine reduce electrical coupling between the axon terminals of the turtle horizontal cells by promoting the release of endogenous dopamine from the dopaminergic amacrine cells previously identified (Witkovsky et al., 1984). Electron-microscopic observations revealed that 6-OH-dopamine selectively attacked this population of amacrine cells. No degenerating terminals were found adjacent to the horizontal cell axon terminals. On this basis, we postulate that dopamine reaches the horizontal cell by diffusion through the extracellular space.