Exploring avian deep-brain photoreceptors and their role in activating the neuroendocrine regulation of gonadal development

Abstract

In the eyes of mammals, specialized photoreceptors called intrinsically photosensitive retinal ganglion cells (ipRGC) have been identified that sense photoperiodic or daylight exposure, providing them over time with seasonal information. Detectors of photoperiods are critical in vertebrates, particularly for timing the onset of reproduction each year. In birds, the eyes do not appear to monitor photoperiodic information; rather, neurons within at least 4 different brain structures have been proposed to function in this capacity. Specialized neurons, called deep brain photoreceptors (DBP), have been found in the septum and 3 hypothalamic areas. Within each of the 4 brain loci, one or more of 3 unique photopigments, including melanopsin, neuropsin, and vertebrate ancient opsin, have been identified. An experiment was designed to characterize electrophysiological responses of neurons proposed to be avian DBP following light stimulation. A second study used immature chicks raised under short-day photoperiods and transferred to long day lengths. Gene expression of photopigments was then determined in 3 septal-hypothalamic regions. Preliminary electrophysiological data obtained from patch-clamping neurons in brain slices have shown that bipolar neurons in the lateral septal organ responded to photostimulation comparable with mammalian ipRGC, particularly by showing depolarization and a delayed, slow response to directed light stimulation. Utilizing real-time reverse-transcription PCR, it was found that all 3 photopigments showed significantly increased gene expression in the septal-hypothalamic regions in chicks on the third day after being transferred to long-day photoperiods. Each dissected region contained structures previously proposed to have DBP. The highly significant increased gene expression for all 3 photopigments on the third, long-day photoperiod in brain regions proposed to contain 4 structures with DBP suggests that all 3 types of DBP (melanopsin, neuropsin, and vertebrate ancient opsin) in more than one neural site in the septal-hypothalamic area are involved in reproductive function. The neural response to light of at least 2 of the proposed DBP in the septal/hypothalamic region resembles the primitive, functional, sensory ipRGC well characterized in mammals.

Keywords: hypothalamus; photopigment; septum; thyroid stimulating hormone β and follicle-stimulating hormone β; thyrotropin-releasing hormone and gonadotropin-releasing hormone-1.

Figure 1.

Sagittal view of the mammalian brain. Intrinsically photosensitive retinal ganglion cells (ipRGC) project to the suprachiasmatic nucleus (SCN), paraventricular nucleus (PVN), and origin of the sympathetic arm of the autonomic nervous system called the intermediolateral nucleus (IML). The IML projects to the superior cervical ganglion (SCG) followed by the pineal gland (PIN). This pathway regulates melatonin release by the pineal and can provide seasonal information to animals (see text). CG, ciliary ganglion; EW, Edinger-Westphal nucleus; IGL, intergeniculate leaflet; LGN_v, ventral division of lateral geniculate nucleus; OPN, olivary pretectal nucleus, structures involved in the pupillary light reflex (modified from Berson, 2003). Reprinted from Trends in Neurosciences, volume 26, D. M. Berson, Strange vision: Ganglion cells as circadian photoreceptors, pages 314–320, copyright 2003, with permission from Elsevier..

Figure 2.

A. Unifying model for pars tuberalis (PT)-dependent photoperiodic regulation of seasonal endocrine function in mammals and birds. Within brackets are deep brain photoreceptors (DBP) found in avian species not mammals. Additionally, T₄ shown in brackets is available to the PT in both mammals and birds via the portal blood supply, cerebrospinal fluid, or both, whereas melatonin influences PT function in mammals (see text). FSH, follicle-stimulating hormone; GnRH, gonadotropin-releasing hormone; LH, luteinizing hormone; PD, pars distalis; PT, pars tuberalis; T₃, 3,5,3′-triiodothyronine; T₄, thyroxine; TSH, thyroid-stimulating hormone; 3V, third ventricle (modified from Hazlerigg and Loudon, 2008). Reprinted from Current Biology, volume 18, D. Hazlerigg and A. Loudon, New insights into ancient seasonal life timers, pages R795–R804, copyright 2008, with permission from Elsevier. B. Septal-hypothalamic area of birds. Four loci have been proposed to contain DBP: (1) lateral septal organ (LSO) including the lateral bed nucleus of the stria terminalis (BSTL), (2) paraventricular nucleus and medial bed nucleus of the stria terminalis complex (PVN/BSTM); (3) premammillary nucleus (PMM), and (4) paraventricular organ (PVO). The 3 nearly rectangular regions show areas dissected for real-time reverse-transcription PCR: region 1 (R1) Sep/Pre/Ant-Hypo, septal preoptic anterior hypothalamic area; region 2 (R2) Mid-Hypo, middle hypothalamic area; region 3 (R3) Post-Hypo, posterior hypothalamic area. AP, anterior pituitary; CO, optic chiasma; GnRH-1, type 1 gonadotropin-releasing hormone; GnRHR, GnRH receptors; ME, median eminence; NHpC, nucleus of the hippocampal commissure; PP, posterior pituitary; TSM, septopallial mesencephalic tract. Color version available in the online PDF.

Figure 3.

A. Light-evoked responses from a neuron in the lateral septal organ, showing slowly rising subthreshold depolarization under current clamp. B. Light-evoked response from another cell under current clamp, showing a light-evoked slow depolarization, which led to delayed suprathreshold spikes. C. Voltage-gated Na currents (shown after leak subtraction) from the same cell as in B, in response to a series of depolarizing voltage steps of increasing amplitude from a holding potential of −70 mV. D. Light-evoked current responses recorded at various holding potentials under voltage clamp, showing a reversal potential near 0 mV. E. Current: voltage (I-V) relation of light evoked currents in D. F. Digital image of an Alexa Fluor 594-filled neuron near the ventricular surface of a septal slice under whole-cell patch showing a bipolar morphology. The intensity of full-field light stimulation was 0.04 nW µm⁻². Color version available in the online PDF.

Figure 4.

A. Adult Japanese quail (solid lines and circles) and adult white-crowned sparrows (dashed lines) were transferred from short to long day lengths starting on d 0. Plasma luteinizing hormone (LH) of Japanese quail significantly increased on d 1 and peaked on d 4 (Follett and Davies, 1975). **P < 0.05; ***P < 0.01. B. Weight of testes of broiler chicks transferred from short day (SD) to long day (LD) and measured on d 3, 7, and 28. Weight of testes was significantly increased on d 7 and 28 in LD birds compared with SD controls (P ≤ 0.05).

Figure 5.

Gene expression of photopigments in the 3 septal/diencephalic regions. A. Opsin 4 (Opn4) and Opsin 5 (Opn5) showed sustained increases in mRNA levels in the septal, preoptic, anterior hypothalamic region (SepPre/Ant-Hypo) following long-day (LD) photoperiods on d 3 and 7. B. Vertebrate ancient opsin (VAOpn) displayed an 8-fold increase in gene expression in mid-hypothalamus (Mid-Hypo) on d 3 of LD photostimulation with low levels of gene expression on d 7 and 28 for both experimental groups. C and D. The Opn4 and Opn5 showed extremely high levels of gene expression, respectively, in the posterior hypothalamus (Post-Hypo) on d 3 of LD photostimulation. Low levels of gene expression were displayed for both Opn4 and Opn5 on d 7 and 28 for both the short-day (SD) and LD groups (unpublished data, S. W. Kang and W. J. Kuenzel). Different letters (a–d) above columns in each figure are significantly different (P < 0.05).

Figure 6.

Neuroendocrine gene expression following transfer of male chicks to long-day (LD) photoperiods. A. Increased mRNA levels of thyrotropin-releasing hormone (TRH) in the middle hypothalamic (Mid-Hypo) region were shown solely on d 3 of LD birds. B. Increased thyroid stimulating hormone β (TSHβ) in anterior pituitary (Pit) was displayed on d 3 and 7 of LD photoperiods. C and D. Gene expression of gonadotropin-releasing hormone-1 (GnRH-1) levels in the septal preoptic anterior hypothalamic (SepPreAnt-Hypo) region and middle hypothalamic (Mid-Hypo) region, respectively, on d 3 and 7 after birds were transferred to LD photoperiods. E and F. Gene expression of follicle-stimulating hormone β (FSHβ) and luteinizing hormone β (LHβ), respectively, in the anterior pituitary of birds exposed to LD photoperiods on d 3 and 7 (unpublished data, S. W. Kang and W. J. Kuenzel). SD = short day. Different letters above each column in a figure are significantly different (P < 0.05).