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First published online November 2, 2007
Journal of Experimental Biology 210, 3970-3978 (2007)
Published by The Company of Biologists 2007
doi: 10.1242/jeb.010645

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Area postrema, a brain circumventricular organ, is the site of antidipsogenic action of circulating atrial natriuretic peptide in eels

Takehiro Tsukada^*, Shigenori Nobata, Susumu Hyodo and Yoshio Takei

Laboratory of Physiology, Ocean Research Institute, The University of Tokyo, Tokyo 164-8639, Japan

View larger version (9K): [in this window] [in a new window]   Fig. 1. Diagram showing mid-sagittal plane of eel brain. The numbers on top (1–6) correspond to cross-sectional planes in Figs 2, 4 and 5. OB, olfactory bulb; Tel, telencephalon; TeO, optic tectum; Cb, cerebellum; rMO, rostral part of medulla oblongata; cMO, caudal part of medulla oblongata; P, pineal gland; SD, saccus dorsalis; PON, preoptic nucleus; SV, saccus vasculosus; RF, reticular formation; GVC, glossopharyngeal-vagal motor complex; NSO, spino-occipital motor nucleus; NCC, commissural nucleus of Cajal; AP, area postrema; II, optic nerve; IV, fourth ventricle.

View larger version (87K): [in this window] [in a new window]   Fig. 2. (A) Localization of major brain nuclei in a cross section taken at the caudal part of medulla oblongata. (B) Sham-operated eel, (C) AP-lesioned eel and (D) eel with lesion outside AP. Dotted areas in C and D indicate areas heat-coagulated by electric cautery. Sections are stained with Haematoxylin. Locations of cross sectional planes are shown in Fig. 1. AP, area postrema; NCC, commissural nucleus of Cajal; GVC, glossopharyngeal-vagal motor complex; NSO, spino-occipital motor nucleus; RF, reticular formation. Scale bar, 200 µm.

View larger version (56K): [in this window] [in a new window]   Fig. 3. Surgical procedure for topical microinjection of kainic acid in seawater (SW) eels. (A) Exposed skull of SW eel (left) and corresponding loci of eel brain (right). The drill hole was made around supraoccipital bone to expose the area postrema (AP). Average size of the AP is approximately 0.8 mm. (B) Experimental setup for injection of 50 nl of kainic acid into the AP of SW eels. A glass capillary was connected to the manipulator to facilitate accurate placement of the capillary at the AP. The capillary was also connected to the microsyringe, through which 50 nl of 4 mmol l–1 kainic acid containing 1% Sky Blue 6B in PBS is injected. (C) Distribution of Chicago Sky Blue 6B in eel brain (left, AP-lesioned eel; right, lesion control). Area of the AP is circled by broken line. TeO, optic tectum; Cb, cerebellum; IV, fourth ventricle.

View larger version (111K): [in this window] [in a new window]   Fig. 4. Immunohistochemical staining of NPR-A in eel brain counterstained with Haematoxylin (left) and corresponding cross-sectional planes (right). (A) Immunoreactive signal in capillaries of whole brain area as represented by the olfactory bulb (OB). (B) Immunoreactive neurons in reticular formation (RF), (C) glossopharyngeal-vagal motor complex (GVC), and (D) area postrema (AP) and commissural nucleus of Cajal (NCC) in the medulla oblongata. Approximate areas photographed are indicated by broken squares drawn in cross sectional planes. Regions indicated by arrowheads are magnified and shown in the boxed areas at the bottom of each photograph. Scale bar, 50 µm (magnified view, 30 µm). Locations of cross sectional planes are shown in Fig. 1. NSO, spino-occipital motor nucleus; IV, fourth ventricle.

View larger version (39K): [in this window] [in a new window]   Fig. 5. Distribution of Trypan Blue in the brain of SW eel (left) and corresponding cross-sectional planes (right). Trypan Blue signal (stained strong red color) was detected in (A) the pineal gland (P) and saccus dorsalis (SD); (B) magnocellular part of preoptic nucleus (PM); (C) area postrema (AP) of medulla oblongata. Approximate areas photographed are indicated by broken squares drawn in cross-sectional planes. Magnified views are shown in the boxed areas at left-bottom of B and C. Locations of cross sectional planes are shown in Fig. 1. PP, parvocellular part of preoptic nucleus; NCC, commissural nucleus of Cajal; GVC, glossopharyngeal-vagal motor complex; NSO, spino-occipital motor nucleus; RF, reticular formation. Scale bars, 20 µm.

View larger version (14K): [in this window] [in a new window]   Fig. 6. (A) Effect of heat-coagulative lesioning by electric cautery on antidipsogenic effect of ANP in seawater eels (N=10 for sham-operated eels, open squares; N=10 for AP-lesioned eels, open circles; N=8 for eels with lesion outside AP, filled circles). (B) Effect of chemical lesioning by kainic acid on antidipsogenic effect of ANP in seawater eels (N=7 for sham-operated eels, open squares; N=7 for AP-lesioned eels, open circles; N=7 for eels with lesion outside AP, filled circles). The values are normalized by the averaged drinking rate for 30 min before ANP injection and expressed as percentages (mean ± s.e.m.). Inhibitory effects of ANP on drinking in AP-lesioned eels are compared with those of sham and lesion controls at each time point (*P<0.05; ANOVA followed by Steel–Dwass test).