inactivation no afterpotential D

DEVELOPMENTAL BIOLOGY

Adult

A 2 kb InaD transcript is detected in wild-type flies, which is absent in the eyes absent mutant that lacks compound eyes. The INAD mRNA is also present in the InaD mutant. Anti-InaD antisera recognize the presence of InaD exclusively in the retina of the eye. The antisera also stain three ocelli, simple eyes located at the apex of the head. InaD is closely associated with rhabdomeres, which are visual organelles consisting of densely packed microvilli (Shieh, 1995).

Effects of Mutation or Deletion

An electroretinogram (ERG) is the extracellular recording of the light-induced electrical activity in the compound eye. A commonly used ERG paradigm to look for mutants with defective vision is to stimulate white-eyed flies with intense light of various wavelengths. In particular, in intense blue light (wavelength 480 nm, which corresponds to the activation wavelength of the major rhodopsin), will convert a substantial amount of rhodopsin into metarhodopsin and bring about a prolonged depolarizing afterpotential (PDA) that persists even after the stimulus has been terminated. During a PDA, photoreceptor cells show a much reduced response to a subsequent pulse of blue light, which is thought to be due to inactivation of photorecepotr cells (when entering a complete PDA, the response to a subsequent pulse of blue light is absent). A PDA can be terminated by a pulse of orange light (wavelength 580 nm), which photoconverts metarhodopsin to rhodopsin. In b rief, the visual physiology of wild-type flies is characterized by the presence of a PDA and inactivation of photoreceptor cells by a pulse of blue light. In contrast, the response of InaD mutant flies to blue light is characterized by the absence of a PDA. Despite not being able to maintain a PDA, the photoreceptors become inactivated and show a much reduced response to a second light stimulus. InaD^p215 is dominant (i.e., both heterozygotes and homozygotes display abnormal phenotype. The InaD^p215 homozyotes exhibit a much more severe phenotype, since the rate of initial decay following the peak responses is greater than that of heterozygotes. Patch-clamp recordings from isolated photoreceptor cells of InaD^p215 show a slow deactivation of the light-induced current. This defective deactivation of InaD appears dependent on calcium influx; removal of extracellular calcium masks its abnormal phenotype. Moreover, InaD photoreceptors show increased sensitivity to dim light. It is proposed that InaD is involved in the negative feedback regulation of the light-activated signaling cascade in Drosophila photoreceptors (Shieh, 1995).

How are signaling molecules organized into different pathways within the same cell? In Drosophila, the inaD gene encodes a protein consisting of five PDZ domains that serves as a scaffold to assemble different components of the phototransduction cascade, including the principal light-activated ion channels, the effector phospholipase C-beta and protein kinase C. Null inaD mutants have a dramatically reorganized subcellular distribution of signaling molecules, and a total loss of transduction complexes. Also, mutants defective in a single PDZ domain produce signaling complexes that lack the target protein and display corresponding defects in their physiology. A picture emerges of a highly organized unit of signaling, a 'transducisome', with PDZ domains functioning as key elements in the organization of transduction complexes in vivo (Tsunoda, 1997).

Flies mutant for the gene coding for the TRP channel have been shown to undergo slow, progressive retinal degeneration. Since TRP is severely mislocalized in InaD^P215 flies, it was asked whether InaD^P215 is also characterized by retinal degeneration. Wild-type flies do not undergo any discernible age-dependent retinal degeneration; therefore, the morphology at 25 days of age is indistinguishable from newly eclosed flies. Young InaD^P215 exhibit a morphology that deviates little if any from wild-type flies. However, the rhabdomeres in inaD flies, aged for 25 days posteclosion under a 12 hr light-12 hr dark cycle, are either missing, reduced in size, or altered in shape. The degeneration in trp^P301 flies, which were also aged for 25 days under a 12 hr light-12 hr dark cycle, is very similar, although the sizes of some InaD^P215 rhabdomeres are slightly larger than trp^P301. Microvill within the affected InaD^P215 rhabdomeres appear swollen and distended, when compared with wild type. Furthermore, a significant amount of vesiculation is observed in the microvilli of the degenerating rhabdomeres. The swollen microvilli and vesiculation of the rhabdomeral membrane observed in 25-day-old ommatidia closely resembles that reported in trp mutant rhabdomeres of similar age. Degeneration of the central R7 rhabdomere is less pronounced than in the R1-R6 cells, again similar to the degeneration seen in trp^P301. As with trp, the degeneration in InaD^P215 is light-dependent, since no degeneration is detected after aging the flies for 25 days in the dark (Chevesich, 1997).

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date revised: 15 April 2007

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