Desensitization and Recovery of Crayfish Photoreceptors Upon Delivery of a Light Stimulus

A method to study desensitization and recovery of crayfish photoreceptors is presented. We performed intracellular electrical recordings of photoreceptor ...

Universidad Nacional Autónoma de México

One of the most important functions modulated by melatonin is the synchronization of circadian rhythms. In crayfish (Procambarus clarkii), we have obtained evidence that the amplitude of the electrical response to light of the retinal photoreceptors the receptor potential, is modified by the action of melatonin and that the magnitude of this action depends on the circadian time of melatonin application. In contrast, the electroretinogram (ERG) circadian rhythm can be synchronized by either single or periodic melatonin application. In this work we hypothesized that, in crayfish, melatonin acts on effectors and on pacemaker of ERG circadian rhythm as a non-photic synchronizer. Melatonin could be a hormone that sends a signal of darkness to the ERG circadian system.

Melatonin modulates the ERG circadian rhythm in crayfish.

Retinular photoreceptors are structures involved in the expression and synchronization of the circadian rhythm of sensitivity to light in crayfish. To determine whether melatonin possesses a differential effect upon the receptor potential (RP) amplitude of retinular photoreceptors circadian time (CT)-dependent, we conducted experiments by means of applying melatonin every 2h during a 24-hour cycle. Melatonin with 100 nM increased RP amplitude during subjective day to a greater degree than during subjective night. To determine whether MT(2) melatonin receptors regulate the melatonin-produced effect, we carried out two experiments, circadian times (CTs) 6 and 18, which included the following: (1) application of the MT(2) receptor selective agonist 8-M-PDOT and antagonist DH97, and (2) the specific binding of [(125)I]-melatonin in eyestalk membranes. The amount of 10 nM of 8-M-PDOT increased RP amplitude in a similar manner to melatonin, and 1 nM DH97 abolished the increase produced by melatonin and 8-M-PDOT. Binding of [(125)I]-melatonin was saturable and specific. Scatchard analysis revealed an affinity constant (K(d)) of 1.1 nM and a total number of binding sites (B(max)) of 6 fmol/mg protein at CT 6, and a K(d) of 1.46 nM and B(max) of 7 fmol/mg protein at CT 18. Our results indicate that melatonin increased RP amplitude of crayfish retinular photoreceptors through MT(2)-like melatonin receptors. These data support the idea that melatonin is a signal of darkness for the circadian system in crayfish retinular cells.

MT2-like melatonin receptor modulates amplitude receptor potential in visual cells of crayfish during a 24-hour cycle.

Visual photoreceptors are structures involved in the expression and synchronization of crayfish circadian rhythm of sensitivity to light (electroretinogram, ERG). Considering the relevant role of Pigment dispersing hormone (PDH) in the invertebrate circadian system organization, we study the effect of this substance on the electrical activity of crayfish visual photoreceptors during the 24-h cycle. The study demonstrates that: (1) PDH affects the electrical response to light of crayfish visual photoreceptor cells in a circadian time-dependent manner. (2) The kinetics of the light-elicited current of crayfish visual photoreceptor cells, as well as the ionic permeability underlying the electrical response to light vary over the 24-h cycle. (3) PDH modifies the kinetics and ionic permeability underlying the light-elicited current of crayfish visual photoreceptor cells in a circadian time-dependent manner.

Effect of pigment dispersing hormone on the electrical activity of crayfish visual photoreceptors during the 24-h cycle.

Mathematical models have been very useful in biological research. From the interaction of biology and mathematics, new problems have emerged that have generated advances in the theory, suggested further experimental work and motivated plausible conjectures. From our perspective, it is absolutely necessary to incorporate modeling tools in the study of circadian rhythms and that without a solid mathematical framework a real understanding of them will not be possible. Our interest is to study the main process underlying the synchronization in the pacemaker of a circadian system: these mechanisms should be conserved in all living beings. Indeed, from an evolutionary perspective, it seems reasonable to assume that either they have a common origin or that they emerge from similar selection circumstances. We propose a general framework to understand the emergence of synchronization as a robust characteristic of some cooperative systems of non-linear coupled oscillators. In a first approximation to the problem we vary the topology of the network and the strength of the interactions among oscillators. In order to study the emergent dynamics, we carried out some numerical computations. The results are consistent with experiments reported in the literature. Finally, we proposed a theoretical framework to study the phenomenon of synchronization in the context of circadian rhythms: the dissipative synchronization of nonautonomous dynamical systems. 

Modeling some properties of circadian rhythms.

In this work, we studied the characteristics of recovery from desensitization of the light-elicited current of crayfish. Applying a two-flash protocol, we found that the first flash triggers a current that activates with a noticeable latency, reaches a peak value, and thereafter decays along a single exponential time course. In comparison with the first-elicited current, the current elicited by the second flash not only presents an expected smaller peak current, depending on the time between flashes, but it also displays a different latency and decay time constant. Recovery of the first flash values of these current parameters depends on the circadian time at which the experiments are conducted, and on the presence of pigment-dispersing hormone. Our data also suggest the existence of distinctive desensitized states, whose induction depends on circadian time and the presence of pigment-dispersing hormone.

Desensitization and recovery of crayfish photoreceptors. Dependency on circadian time, and pigment-dispersing hormone.

Melatonin (MEL) is a conserved molecule with respect to its synthesis pathway and functions. In crayfish, MEL content in eyestalks (Ey) increases at night under the photoperiod, and this indoleamine synchronizes the circadian rhythm of electroretinogram amplitude, which is expressed by retinas and controlled by the cerebroid ganglion (CG). The aim of this study was to determine whether MEL content in eyestalks and CG or circulating MEL in hemolymph (He) follows a circadian rhythm under a free-running condition; in addition, it was tested whether MEL might directly influence the spontaneous electrical activity of the CG. Crayfish were maintained under constant darkness and temperature, a condition suitable for studying the intrinsic properties of circadian systems. MEL was quantified in samples obtained from He, Ey, and CG by means of an enzyme-linked immunosorbent assay, and the effect of exogenous MEL on CG spontaneous activity was evaluated by electrophysiological recording. Variation of MEL content in He, Ey, and CG followed a circadian rhythm that peaked at the same circadian time (CT). In addition, a single dose of MEL injected into the crayfish at different CTs reduced the level of spontaneous electrical activity in the CG. Results suggest that the circadian increase in MEL content directly affects the CG, reducing its spontaneous electrical activity, and that MEL might act as a periodical signal to reinforce the organization of the circadian system in crayfish.

Circadian rhythm in melatonin release as a mechanism to reinforce the temporal organization of the circadian system in crayfish.

Melatonin (MEL) is an ancient molecule, broadly distributed in nature from unicellular to multicellular species. MEL is an indoleamine that acts on a wide variety of cellular targets regulating different physiological functions. This review is focused on the role played by this molecule in the regulation of the circadian rhythms in crayfish. In these species, information about internal and external time progression might be transmitted by the periodical release of MEL and other endocrine signals acting through the pacemaker. We describe documented and original evidence in support of this hypothesis that also suggests that the rhythmic release of MEL contributes to the reinforcement of the temporal organization of nocturnal or diurnal circadian oscillators. Finally, we discuss how MEL might coordinate functions that converge in the performance of complex behaviors, such as the agonistic responses to establish social dominance status in  and the burrowing behavior in the secondary digging crayfish 

Beatriz Fuentes-Pardo

Departamento de Fisiología,
Facultad de Medicina,
Departamento de Fisiología, Facultad de Medicina

Universidad Nacional Autónoma de México

Desensitization and Recovery of Crayfish Photoreceptors Upon Delivery of a Light Stimulus

Beatriz Fuentes-Pardo

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Departamento de Fisiología,Facultad de Medicina,Departamento de Fisiología, Facultad de Medicina

Universidad Nacional Autónoma de México

Desensitization and Recovery of Crayfish Photoreceptors Upon Delivery of a Light Stimulus

Departamento de Fisiología,
Facultad de Medicina,
Departamento de Fisiología, Facultad de Medicina