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Adjustments in cholinergic, adrenergic and purinergic control of cardiovascular function in snapping turtle embryos (Chelydra serpentina) incubated in chronic hypoxia

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Abstract

Adenosine is an endogenous nucleoside that acts via G-protein coupled receptors. In vertebrates, arterial or venous adenosine injection causes a rapid and large bradycardia through atrioventricular node block, a response mediated by adenosine receptors that inhibit adenylate cyclase and decrease cyclic AMP concentration. Chronic developmental hypoxia has been shown to alter cardioregulatory mechanisms in reptile embryos, but adenosine’s role in mediating these responses is not known. We incubated snapping turtle embryos under chronic normoxic (N21; 21 % O2) or chronic hypoxic conditions (H10; 10 % O2) beginning at 20 % of embryonic incubation. H10 embryos at 90 % of incubation were hypotensive relative to N21 embryos in both normoxic and hypoxic conditions. Hypoxia caused a hypotensive bradycardia in both N21 and H10 embryos during the initial 30 min of exposure; however, f H and P m both trended towards increasing during the subsequent 30 min, and H10 embryos were tachycardic relative to N21 embryos in hypoxia. Following serial ≥1 h exposure to normoxic and hypoxic conditions, a single injection of adenosine (1 mg kg−1) was given. N21 and H10 embryos responded to adenosine injection with a rapid and large hypotensive bradycardia in both normoxia and hypoxia. Gene expression for adenosine receptors were quantified in cardiac tissue, and Adora1 mRNA was the predominant receptor subtype with transcript levels 30–82-fold higher than Adora2A or Adora2B. At 70 % of incubation, H10 embryos had lower Adora1 and Adora2B expression compared to N21 embryos. Expression of Adora1 and Adora2B decreased in N21 embryos during development and did not differ from H10 embryos at 90 % of incubation. Similar to previous results in normoxia, H10 embryos in hypoxia were chronically tachycardic compared to N21 embryos before and after complete cholinergic and adrenergic blockade. Chronic hypoxia altered the development of normal cholinergic and adrenergic tone, as well as adenosine receptor mRNA levels. This study demonstrates that adenosine may be a major regulator of heart rate in developing snapping turtle embryos, and that chronic hypoxic incubation alters the response to hypoxic exposure.

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Abbreviations

CAM:

Chorioallantoic membrane

f H :

Heart rate (beats min−1), ‘frequency of heart beats’

H10:

Turtle embryos artificially incubated in chronic hypoxia (10 % O2), beginning at 20 % of incubation/development

N21:

Turtle embryos artificially incubated in chronic normoxia (21 % O2), beginning at 20 % of incubation/development

P m :

Mean arterial pressure (kPa) measured through a CAM artery

70 %:

70 % of embryonic incubation/development for turtle embryos (~55 day total incubation)

90 %:

90 % of embryonic incubation/development for turtle embryos (~55 day total incubation)

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Acknowledgments

The authors sincerely thank the State of Minnesota Department of Natural Resources for special permit No. 18337 to collect turtle eggs. Thanks to Kevin Tate and Kathryn Gruchalla for assistance. This work was supported by NSF CAREER award IBN IOS-0845741 to DAC and NSF award IBN IOS-0923300 to TR. No conflicts of interests, financial or otherwise, are declared by the authors.

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Authors and Affiliations

  1. Department of Biology, McMaster University, Hamilton, ON, Canada

    John Eme

  2. Department of Biology, University of North Dakota, Grand Forks, ND, USA

    Turk Rhen

  3. Department of Biological Sciences, University of North Texas, Denton, TX, USA

    Dane A. Crossley II

Authors
  1. John Eme
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  2. Turk Rhen
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  3. Dane A. Crossley II
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Correspondence to Dane A. Crossley II.

Additional information

Communicated by H.V. Carey.

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Supplementary Figs. S1–S7

S1 shows a neighbor-joining phylogenetic tree comparing amino acid sequences of C. serpentina to two turtles, alligator, chicken, green anole, Tasmanian devil, mouse and human. The scale bar on S1 indicates the number of amino acid changes per site (the number of changes divided by sequence length), and the blackened sequences represent the seven transmembrane domains. S2 and S3 show amino acid and nucleotide alignments for Adora1 receptor, respectively, S4 and S5 show amino acid and nucleotide alignments for Adora2A receptor, respectively, and S6 and S7 show amino acid and nucleotide alignments for Adora2B receptor, respectively (PDF 1,174 kb)

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Eme, J., Rhen, T. & Crossley, D.A. Adjustments in cholinergic, adrenergic and purinergic control of cardiovascular function in snapping turtle embryos (Chelydra serpentina) incubated in chronic hypoxia. J Comp Physiol B 184, 891–902 (2014). https://doi.org/10.1007/s00360-014-0848-6

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  • DOI: https://doi.org/10.1007/s00360-014-0848-6

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