Endothelians are a family of three peptide hormones (ET-1, ET-2, ET-3) with cardiovascular functions that are mediated by two main receptor subtypes,ETA and ETB. In mammals, ETA is responsible for vasoconstriction in blood vessels while ETB is believed to facilitate their relaxation. Endothelians have also been described in some fish species and play a role in gill vascular resistance, dorsal aortic blood pressure and the contraction of pillar cells, which are interspersed throughout the gill lamellae and credited with keeping the `roof' and `floor'of the lamellae together. Ultimately, these vascular effects of endothelians have direct implications on gill blood flow and gas exchange. Furthermore,endothelian-like immunoreactivity has been observed in the gill neuroepithelial cells. Although the function and cellular localization of endothelians are relatively well described in fish, information on the receptors that mediate its response is contradictory. And so, Kåre-Olav Stensløkken, Lena Sundin and Göran Nilsson set out to clarify the nature of endothelian receptors in fish gills in an onslaught of experimentation using a combination of cardiovascular physiology, receptor pharmacology and microscopy.
Atlantic cod, mackerel and sculpin were anesthetized and surgically implanted with a variety of catheters to measure different cardiovascular parameters and to permit the injection of various pharmacological agents. To observe gill blood circulation, a digital video camcorder was hooked up to an epi-illumination microscope and placed in close proximity to the fish gill. Using this setup, the team could observe `real-time' changes in the diameter of arteries and pillar cells and make direct measurements of their contraction in response to various compounds.
Stensløkken and the group found that intra-arterial injection of ET-1 resulted in the contraction of gill pillar cells, and a dose-dependent increase in ventral aortic blood pressure and gill vascular resistance of all three fish species, with the cod being the most sensitive. Wondering which type of endothelian receptor mediates pillar cell contraction, they injected the fish with the specific ETB agonist BQ-3020 and found that it mimicked the contractile effect of ET-1 on pillar cells. By contrast,injection of the ETA antagonist BQ-610 did not block the pillar cell contraction induced by ET-1, suggesting that it is an ETB-like receptor and not an ETA receptor that mediates pillar cell contraction. Using antibodies specific for both receptor subtypes, the team revealed ETB-like immunoreactivity in the gill lamellae adjacent to the pillar cells, consistent with the receptor's role in pillar cell contractility in neuroepithelial cells and blood vessels. By contrast, very little ETA-like immunoreactivity was detected in the lamellar region, although very faint ETA-like staining was found adjacent to pillar cells in some lamellae, suggesting that ETA's role in pillar cell contraction is minor at best. Instead, ETA was clearly detected in gill nerve fibers, suggesting that endothelian and ETAmight be involved with the transmission of sensory signals and not vasoconstriction.
With an arsenal of techniques and a thorough approach, Stensløkken,Sundin and Nilsson have been successful in their quest to further characterize endothelian mechanisms in fish and relieve some of the confusion about the receptors that mediate the hormones' effects. There are still many questions left unanswered; however, we now know that, in general, fish pillar cells are sensitive to ET-1. Furthermore, at least in the Atlantic cod, the contractile effects of ET-1 are mediated by ETB-like receptors, while ETA-like receptors may serve another, as yet unknown, function.