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PACAP-deficient mice exhibit light parameter-dependent abnormalities on nonvisual photoreception and early activity onset.

Kawaguchi C, Isojima Y, Shintani N, Hatanaka M, Guo X, Okumura N, Nagai K, Hashimoto H, Baba A - PLoS ONE (2010)

Bottom Line: The photopigment melanopsin has been suggested to act as a dominant photoreceptor in nonvisual photoreception including resetting of the circadian clock (entrainment), direct tuning or masking of vital status (activity, sleep/wake cycles, etc.), and the pupillary light reflex (PLR).These data suggest that the dysfunctions of entrainment and masking were caused by the loss of PACAP, not by the loss of light input itself.These results indicate that PACAP regulates particular nonvisual light responses by conveying parametric light information--that is, intensity and duration.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.

ABSTRACT

Background: The photopigment melanopsin has been suggested to act as a dominant photoreceptor in nonvisual photoreception including resetting of the circadian clock (entrainment), direct tuning or masking of vital status (activity, sleep/wake cycles, etc.), and the pupillary light reflex (PLR). Pituitary adenylate cyclase-activating polypeptide (PACAP) is exclusively coexpressed with melanopsin in a small subset of retinal ganglion cells and is predicted to be involved extensively in these responses; however, there were inconsistencies in the previous reports, and its functional role has not been well understood.

Methodology/principal findings: Here we show that PACAP-deficient mice exhibited severe dysfunctions of entrainment in a time-dependent manner. The abnormalities in the mutant mice were intensity-dependent in phase delay and duration-dependent in phase advance. The knockout mice also displayed blunted masking, which was dependent on lighting conditions, but not completely lost. The dysfunctions of masking in the mutant mice were recovered by infusion of PACAP-38. By contrast, these mutant mice show a normal PLR. We examined the retinal morphology and innervations in the mutant mice, and no apparent changes were observed in melanopsin-immunoreactive cells. These data suggest that the dysfunctions of entrainment and masking were caused by the loss of PACAP, not by the loss of light input itself. Moreover, PACAP-deficient mice express an unusually early onset of activities, from approximately four hours before the dark period, without influencing the phase of the endogenous circadian clock.

Conclusions/significance: Although some groups including us reported the abnormalities in photic entrainments in PACAP- and PAC(1)-knockout mice, there were inconsistencies in their results. The time-dependent dysfunctions of photic entrainment in the PACAP-knockout mice described in this paper can integrate the incompatible data in previous reports. The recovery of impaired masking by infusion of PACAP-38 in the mutant mice is the first direct evidence of the relationship between PACAP and masking. These results indicate that PACAP regulates particular nonvisual light responses by conveying parametric light information--that is, intensity and duration. The "early-bird" phenotype in the mutant mice originally reported in this paper supposed that PACAP also has a critical role in daily behavioral patterns, especially during the light-to-dark transition period.

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Retinal morphology, innervation and expression of PACAP-related genes in the eye.(A) Immunohistochemical analysis of melanopsin localization (brown) in the retina. Slices were counterstained with hematoxylin. GCL: ganglion cell layer; IPL: inner plexiform layer; INL: inner nuclear layer; OPL: outer plexiform layer; ONL: outer nuclear layer; RPE: retinal pigment epithelium. Scale bar = 10 µm. (B) Mean cell number in the GCL (n = 4). (C) Percentage of melanopsin-expressing cells in the GCL (n = 4). (D) Bright-field photomicrographs of retinal innervation to the SCN and the OPT, stained purple. (E) Expression levels of PACAP-related genes in the whole eye (n = 3–4). Means ± SEM are presented.
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pone-0009286-g004: Retinal morphology, innervation and expression of PACAP-related genes in the eye.(A) Immunohistochemical analysis of melanopsin localization (brown) in the retina. Slices were counterstained with hematoxylin. GCL: ganglion cell layer; IPL: inner plexiform layer; INL: inner nuclear layer; OPL: outer plexiform layer; ONL: outer nuclear layer; RPE: retinal pigment epithelium. Scale bar = 10 µm. (B) Mean cell number in the GCL (n = 4). (C) Percentage of melanopsin-expressing cells in the GCL (n = 4). (D) Bright-field photomicrographs of retinal innervation to the SCN and the OPT, stained purple. (E) Expression levels of PACAP-related genes in the whole eye (n = 3–4). Means ± SEM are presented.

Mentions: PACAP exerts neurotrophic and neuroprotective effects and modulates cellular differentiation and survival [12]. In the retina, PACAP promotes retinal development through its specific receptor, PAC1 [22], [23]. Thus, we examined whether there was any remodeling in terms of morphology, neuronal projections and PACAP-related gene expression, in the retinas of Adcyap1−/− mice. Adcyap1−/− mice showed retention of somatic and dendritic/axonal layers, and melanopsin expression in the ganglion cell layer (Fig. 4A–C). Anterograde tracing with the use of horseradish peroxidase–labeled wheat germ agglutinin disclosed normal projections from RGCs to the SCN or OPT in the mutants (Fig. 4D). Gene expression analysis (Fig. 4E) indicated that there were no changes in the expression of VIP (a peptide that has approximately 70% amino acid sequence identity with PACAP), VPAC2 (VIP receptor subtype 2: a receptor for both PACAP and VIP), and Thy1 (thymus cell antigen 1, theta: a marker of the retinal ganglion cell layer) [24]. PAC1 expression showed a tendency to increase in Adcyap1−/− mice, but this difference did not reach statistical significance (P = 0.10). We confirmed the complete loss of PACAP mRNA in Adcyap1−/− eye.


PACAP-deficient mice exhibit light parameter-dependent abnormalities on nonvisual photoreception and early activity onset.

Kawaguchi C, Isojima Y, Shintani N, Hatanaka M, Guo X, Okumura N, Nagai K, Hashimoto H, Baba A - PLoS ONE (2010)

Retinal morphology, innervation and expression of PACAP-related genes in the eye.(A) Immunohistochemical analysis of melanopsin localization (brown) in the retina. Slices were counterstained with hematoxylin. GCL: ganglion cell layer; IPL: inner plexiform layer; INL: inner nuclear layer; OPL: outer plexiform layer; ONL: outer nuclear layer; RPE: retinal pigment epithelium. Scale bar = 10 µm. (B) Mean cell number in the GCL (n = 4). (C) Percentage of melanopsin-expressing cells in the GCL (n = 4). (D) Bright-field photomicrographs of retinal innervation to the SCN and the OPT, stained purple. (E) Expression levels of PACAP-related genes in the whole eye (n = 3–4). Means ± SEM are presented.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2823792&req=5

pone-0009286-g004: Retinal morphology, innervation and expression of PACAP-related genes in the eye.(A) Immunohistochemical analysis of melanopsin localization (brown) in the retina. Slices were counterstained with hematoxylin. GCL: ganglion cell layer; IPL: inner plexiform layer; INL: inner nuclear layer; OPL: outer plexiform layer; ONL: outer nuclear layer; RPE: retinal pigment epithelium. Scale bar = 10 µm. (B) Mean cell number in the GCL (n = 4). (C) Percentage of melanopsin-expressing cells in the GCL (n = 4). (D) Bright-field photomicrographs of retinal innervation to the SCN and the OPT, stained purple. (E) Expression levels of PACAP-related genes in the whole eye (n = 3–4). Means ± SEM are presented.
Mentions: PACAP exerts neurotrophic and neuroprotective effects and modulates cellular differentiation and survival [12]. In the retina, PACAP promotes retinal development through its specific receptor, PAC1 [22], [23]. Thus, we examined whether there was any remodeling in terms of morphology, neuronal projections and PACAP-related gene expression, in the retinas of Adcyap1−/− mice. Adcyap1−/− mice showed retention of somatic and dendritic/axonal layers, and melanopsin expression in the ganglion cell layer (Fig. 4A–C). Anterograde tracing with the use of horseradish peroxidase–labeled wheat germ agglutinin disclosed normal projections from RGCs to the SCN or OPT in the mutants (Fig. 4D). Gene expression analysis (Fig. 4E) indicated that there were no changes in the expression of VIP (a peptide that has approximately 70% amino acid sequence identity with PACAP), VPAC2 (VIP receptor subtype 2: a receptor for both PACAP and VIP), and Thy1 (thymus cell antigen 1, theta: a marker of the retinal ganglion cell layer) [24]. PAC1 expression showed a tendency to increase in Adcyap1−/− mice, but this difference did not reach statistical significance (P = 0.10). We confirmed the complete loss of PACAP mRNA in Adcyap1−/− eye.

Bottom Line: The photopigment melanopsin has been suggested to act as a dominant photoreceptor in nonvisual photoreception including resetting of the circadian clock (entrainment), direct tuning or masking of vital status (activity, sleep/wake cycles, etc.), and the pupillary light reflex (PLR).These data suggest that the dysfunctions of entrainment and masking were caused by the loss of PACAP, not by the loss of light input itself.These results indicate that PACAP regulates particular nonvisual light responses by conveying parametric light information--that is, intensity and duration.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.

ABSTRACT

Background: The photopigment melanopsin has been suggested to act as a dominant photoreceptor in nonvisual photoreception including resetting of the circadian clock (entrainment), direct tuning or masking of vital status (activity, sleep/wake cycles, etc.), and the pupillary light reflex (PLR). Pituitary adenylate cyclase-activating polypeptide (PACAP) is exclusively coexpressed with melanopsin in a small subset of retinal ganglion cells and is predicted to be involved extensively in these responses; however, there were inconsistencies in the previous reports, and its functional role has not been well understood.

Methodology/principal findings: Here we show that PACAP-deficient mice exhibited severe dysfunctions of entrainment in a time-dependent manner. The abnormalities in the mutant mice were intensity-dependent in phase delay and duration-dependent in phase advance. The knockout mice also displayed blunted masking, which was dependent on lighting conditions, but not completely lost. The dysfunctions of masking in the mutant mice were recovered by infusion of PACAP-38. By contrast, these mutant mice show a normal PLR. We examined the retinal morphology and innervations in the mutant mice, and no apparent changes were observed in melanopsin-immunoreactive cells. These data suggest that the dysfunctions of entrainment and masking were caused by the loss of PACAP, not by the loss of light input itself. Moreover, PACAP-deficient mice express an unusually early onset of activities, from approximately four hours before the dark period, without influencing the phase of the endogenous circadian clock.

Conclusions/significance: Although some groups including us reported the abnormalities in photic entrainments in PACAP- and PAC(1)-knockout mice, there were inconsistencies in their results. The time-dependent dysfunctions of photic entrainment in the PACAP-knockout mice described in this paper can integrate the incompatible data in previous reports. The recovery of impaired masking by infusion of PACAP-38 in the mutant mice is the first direct evidence of the relationship between PACAP and masking. These results indicate that PACAP regulates particular nonvisual light responses by conveying parametric light information--that is, intensity and duration. The "early-bird" phenotype in the mutant mice originally reported in this paper supposed that PACAP also has a critical role in daily behavioral patterns, especially during the light-to-dark transition period.

Show MeSH
Related in: MedlinePlus