A blind circadian clock in cavefish reveals that opsins mediate peripheral clock photoreception.

The circadian clock is synchronized with the day-night cycle primarily by light. Fish represent fascinating models for deciphering the light input pathway to the vertebrate clock since fish cell clocks are regulated by direct light exposure. Here we have performed a comparative, functional analysis of the circadian clock involving the zebrafish that is normally exposed to the day-night cycle and a cavefish species that has evolved in perpetual darkness. Our results reveal that the cavefish retains a food-entrainable clock that oscillates with an infradian period. Importantly, however, this clock is not regulated by light. This comparative study pinpoints the two extra-retinal photoreceptors Melanopsin (Opn4m2) and TMT-opsin as essential upstream elements of the peripheral clock light input pathway.

Encephalic photoreception and phototactic response in the troglobiont Somalian blind cavefish Phreatichthys andruzzii.

Many physiological and behavioural responses to changes in environmental lighting conditions are mediated by extraocular photoreceptors. Here we investigate encephalic photoreception in Phreatichthys andruzzii, a typical cave-dwelling fish showing an extreme phenotype with complete anophthalmy and a reduction in size of associated brain structures. We firstly identified two P. andruzzii photopigments, orthologues of rod opsin and exo-rod opsin. In vitro, both opsins serve as light-absorbing photopigments with λ(max) around 500 nm when reconstituted with an A(1) chromophore. When corrected for the summed absorption from the skin and skull, the spectral sensitivity profiles shifted to longer wavelengths (rod opsin: 521 nm; exo-rod opsin: 520 nm). We next explored the involvement of both opsins in the negative phototaxis reported for this species. A comparison of the spectral sensitivity of the photophobic response with the putative A(2) absorbance spectra corrected for skin/skull absorbance indicates that the A(2) versions of either or both of these pigments could explain the observed behavioural spectral sensitivity.

Ultrastructure and distribution of superficial neuromasts of blind cavefish, Phreatichthys andruzzii, juveniles.

Transmission and scanning electron microscopy (TEM, SEM) were used to study the ultrastructure of superficial neuromasts in 15 six-month old blind cavefish juveniles, Phreatichthys andruzzii (Cyprinidae). In five specimens examined with SEM, the number of superficial neuromasts over the fish body (480-538) was recorded. They were localized mainly on the head (362-410), including the dorsal surface, the mentomandibular region, and laterally from the mouth to the posterior edge of the operculum. Neuromasts were also present laterally on the trunk and near the caudal fin (116-140). A significantly higher number of neuromasts were present on the head compared to the trunk (t-test, P < 0.05). Superficial neuromasts of the head and those along the trunk were similar in ultrastructure. Each neuromast comprised sensory hair cells surrounded by nonsensory support cells (mantle cells and supporting basal cells) with the whole covered by a cupula. Each hair cell was pear-shaped, 15-21 microm high and 4-6 microm in diameter, with a single long kinocilium and several short stereocilia. Most support cells were elongated, with nuclei occupying a large portion of the cytoplasm. In the margin of the neuromast, mantle cells were particularly narrow. Both types of support cells had well-developed Golgi apparatus and rough endoplasmic reticulum. The number of hair cells and nonsensory support cells of the anterior lateral line (head) did not differ significantly from those of the posterior lateral line (trunk) (t-test, P > 0.05).

The lateral line system in larvae of the blind cyprinid cavefish, Phreatichthys andruzzii.

Scanning and transmission electron microscopic observations were made on the anterior and posterior lateral line system of larvae of the blind cavefish, Phreatichthys andruzzii. The investigations were conducted on 36 specimens belonging to three different ages of juvenile: 7, 14, and 28 days posthatching. The basic sensory unit of the mechanoreceptive system is a superficial "neuromast," which is composed of sensory hair cells (HCs) embedded in nonsensory support cells (SCs) frequently covered with a cupula. Head neuromasts had the longest cupulae. These organs presented the same structure and ultrastructure in the head and along the body of all specimens of P. andruzzii of different ages. Hair cells of neuromasts of P. andruzzii larvae were pear-shaped cells approximately 10-20 microm high and 3-5 microm in diameter. The nucleus of each hair cell was large and basally placed, the cytoplasm contained numerous mitochondria and each HC had a single long kinocilium and several short stereocilia. The majority of SCs were elongated in shape and their nuclei occupied a significant portion of the cytoplasm. The support cells at the margin of the neuromast were particularly thin. Both types of support cells possessed well-developed rough endoplasmic reticulum and Golgi apparatus. The size and number of neuromasts and their component cells increased with the age of larvae (ANOVA, P < 0.05). The dimensions of the neuromasts of the anterior lateral line (head) did not differ significantly from those of the posterior lateral line (trunk) (ANOVA, P > 0.05).

Terapêutica fotodinâmica com ácido delta-aminole-vulínico em neoplasias queratinocíticas superficiais / Photodynamic therapy with delta-aminolevulinic acid for superficial keratinocytic neoplasms

A terapêutica fotodinâmica (TFD) é técnica de tratamento em que são aplicadas, nos tecidos, substâncias fotossensibilizantes, posteriormente ativadas com luzes de comprimentos de onda específicos, com a finalidade de produzir destruição celular, por meio da ação de produtos citotóxicos fotoativados. Tal método tem sido utilizado em diversos tipos de neoplasias cutâneas, com resultados bastante entusiasmadores. Avaliar o efeito de uma única sessão de terapêutica fotodinâmica (TFD) com ácido delta aminolevulínico (ALA) tópico, estimulado por luz não coerente, em ceratoses actínicas (CAC), carcinomas basocelulares (CBC) superficiais e doenças de Bowen (DB). Utilizando uma fonte de luz não coerente, emitida por protótipo desenvolvido no Serviço de Engenharia Biomédica do Hospital de Clínicas de Porto Alegre (HCPA), para ativar o ácido delta-aminolevulínico (ALA) a 20 por cento, aplicado com oclusão, foram tradadas 80 lesões cutâneas, em 52 pacientes. Entre as lesões, 32 (40 por cento) eram carcinomas basocelulares superficiais, 37 (46,3 por cento) ceratoses actinícas e 11 (13,7 por cento) doenças de Bowen. Em 23 casos (28,7 por cento), além das características clínicas, o diagnóstico foi confirmado por biópsia e exame histopatológico. Não foi possível obter o seguimento de cinco lesões tratadas. Das 75 lesões que puderam ser avaliadas, 41 (54,6 por cento) apresentaram cura, 22 (29,4 por cento) apresentaram melhora, e 12 (16,0 por cento) permaneceram inalteradas, tendo o tratamento sido considerado ineficaz nestas últimas. Das 35 ceratoses actínicas, 23 (65,7 por cento) obtiveram cura, 7 (20 por cento) melhoraram, e cinco (14,3 por cento) não responderam ao tratamento. Trinta carcinomas basocelulares superficiais foram tratados, com cura em 10 (33,3 por cento), melhora em 13 (43,4 por cento) e sem alterações em sete (23,3 por cento). Dos 10 casos de doença de Bowen avaliados, oito (80 por cento) curaram e dois (20 por cento) melhoraram. Os efeitos colaterais observados foram ardência em 24 lesões (32 por cento), dor em cinco (6,7 por cento) e prurido em um caso (1,3 por cento. Os resultados obtidos no trabalho assemelham-se, em parte, aos relatados na literatura com aplicação única da terapêutica fotodinâmica com ácido delta-aminolevulínico em neoplasias queratinocíticas superficiais, principalmente nas ceratoses actínicas e na doença de Bowen