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COMPARATIVE AND EVOLUTIONARY PHYSIOLOGY
Department of Zoology, University of Gothenburg, Gothenburg, Sweden
Submitted 25 June 2008 ; accepted in final form 21 October 2008
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ABSTRACT |
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 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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-adrenoceptor-mediated increase in the dorsal aortic blood pressure (Pda), with no change in cardiac output (CO), gut blood flow (Qcma), or heart rate. To investigate if feeding habits affect these patterns and to compare the differences between gastric distension alone and feeding in the same experimental setting, we used the short-horn sculpin (Myoxocephalus scorpius), an inactive ambush predator with a capacity to eat large meals. An inflatable balloon was placed in the stomach of one group while another group was fed fish meat. When distending the stomach with a volume corresponding to a meal of 8–10% body weight, there is a profound and long-lasting increase in systemic (123 ± 27%) and gastrointestinal (Rcma; 82 ± 24%) vascular resistance, leading to an increase in Pda (19%) without any change in CO or Qcma. After force-feeding, there is a rapid transient increase in Rcma (24 ± 4%) and an even larger Pda response (53%). There is also a subsequent increase in both CO (28 ± 8%) and Qcma (27 ± 9%) after 30 min. By 15 h, CO and Qcma increase further (41 ± 11 and 63 ± 14%, respectively), and this increase persists for up to 60 h. The increase in Qcma is mediated via both an increase in CO and a shunting of blood from the systemic circulation via a decrease in Rcma (34 ± 7%). In conclusion, the response to mechanical distension of the stomach is similar to what we have described in rainbow trout, and the postprandial gastrointestinal hyperemia is most likely chemically induced. postprandial blood flow; coeliaco-mesenteric artery; mechanical stimuli; shunting of blood; pressor response; vascular resistance
The increase in the gastrointestinal blood flow depends on either an increased cardiac output (CO; see Refs. 2, 4, 5, 22, 33), a shunting of blood from less active tissue to the gastrointestinal tract (16, 41, 42), or a combination of the two (18). In contrast to mammals, fish show a larger increase in CO that in some cases might account for the entire increase in gut blood flow (4, 5).
However, the information is scarce regarding to what extent the mechanical distension of the stomach following feeding might contribute to the cardiovascular response. Traditionally, it is believed that the majority of this response is due to the chemical stimuli from components of the hydrolyzed food such as glucose and fatty acids (6, 8, 10, 11, 13, 37) as well as gastrointestinal secretions (24, 25). The possible regulation behind a nutrient-induced hyperemia in mammals has been reviewed extensively (19, 20, 29). However, few studies have considered the importance of mechanical distension of the stomach (26–28) and intestine (11) in the regulation of gut blood flow in mammals.
We recently published the first study on the effects of mechanical stomach distension on the gastrointestinal blood flow in rainbow trout (Oncorhynchus mykiss) (36). When the stomach was mechanically distended using an inflatable nitrile balloon, the dorsal aortic pressure (Pda) increased by up to 29% within minutes after filling the balloon with a volume corresponding to the natural meal/prey size (2% of body wt) (23, 32). This increase was mediated via an increase in the systemic circulation through the activation of -adrenoceptors. No gastrointestinal hyperemia was seen, and we speculate that additional stimuli are needed to evoke an increased gut blood flow and that the increased systemic resistance prepares for the shunting of blood to the gut. This corresponds well with the response seen in mammals during the anticipation and ingestion phase (18).
To further establish the significance of mechanical stimuli in the postprandial gut blood flow response seen in fish (2–5, 38), we compared the response to mechanical stimuli (gastric distension) with the effects when feeding the fish a regular fish diet under the same experimental conditions. By selecting the short-horn sculpin (Myoxocephalus scorpius), an inactive ambush feeder, we were also able to investigate how different feeding habits affect the response. In contrast to the rainbow trout, this species is able to swallow larger quantities of food in a single meal, and an average-sized meal is considered to be 8–10% body weight (observation from freely feeding fish). This would lead to a larger mechanical stimuli, and possibly a more rapid and profound cardiovascular response.
Furthermore, we also characterized the central cardiovascular system of the short-horn sculpin using corrosion casts as previously described by Murakami (30, 31). This enabled us to correctly place the gastrointestinal flow probe on the coeliaco-mesenteric artery (CMA).
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MATERIALS AND METHODS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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Corrosion cast of the arterial vascular system.
To reveal the vasculature of the gastrointestinal system and the major vessels of the systemic circulation, as shown in Fig. 1, we used a corrosion casting technique described by Murakami (30). Short-horn sculpins (n = 3; 
200–250 g) were anesthetized using a high dose of MS-222 (300 mg/l) in seawater. A catheter (PE-90; polyethylene), filled with heparinized saline (100 IU/ml; 0.09%), was placed in the ventral aorta just anterior to the heart, and the fish was killed by exsanguination. The entire vasculature was then flushed with ample quantities of heparinized saline (100 IU/ml; 0.09%) followed by saline containing sodium nitroprusside (6 g/l) too dilate the vessels. The vascular system was then filled with the Mercox CL-2R casting solution (Ladd Research). The solution had been prepared at a resin-to-catalyst ratio of 20:1 (20–30 cP at 25°C). Once the vascular system was completely filled, the fish was placed in warm water to enhance polymerization. When complete polymerization was achieved, the fish was placed in a saturated solution of KOH to remove tissues and reveal the cast. After 48 h, most of the tissues had been fragmented or saponified, and the remaining bones were removed by hand along with some of the cast to expose only the major vessels. Photos (Canon Digital Ixus 400) were taken to document the vasculature structure revealed by the cast.
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Measuring the gastric emptying time.
To establish the rate in which the fish empty their stomach after a meal, we fed (freely feeding) short-horn sculpins (n = 16) with fillets of Hoki corresponding to 8–10% of body weight. The fish had been fasted for at least 6 days and accustomed to the experimental chamber for 24 h before the meal. Fish were killed at 5, 24, 48, and 72 h postfeeding, and the stomach contents were carefully removed and weighed. To remove any ambiguity due to differences in the relative water content, the stomach content was desiccated for 24 h at 80°C to remove all water. The values at 5, 24, 48, and 72 h were then compared with the desiccated prefeeding weight set to 100% (Fig. 2).
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1 wk before surgery. Fish were anesthetized in seawater containing MS-222 (100 mg/l) and transferred to the operating table where they were placed on wet rubber foam. Anesthesia was maintained by irrigating the gills with circulating seawater (10°C) containing MS-222 (50 mg/l). To measure the Pda, a PE-50 catheter fitted with a PE-10 catheter at the tip and filled with heparinized saline (100 IU/ml; 0.09%) was inserted in the third EBA of the left side and secured to the skin by a stitch. Venous pressure (Pven) was measured by placing a PE-50 catheter, filled with heparinized saline, in the most ventral of the 4-7 hepatic portal veins through an incision made ventrolaterally on the left side just behind the pectoral fin. Through the same incision, a Doppler flow probe was placed on the CMA just before its bifurcation (Fig. 1). CO was measured by placing a Doppler flow probe on the ventral aorta through an incision in the ventral midline where the two operculas meet. Both flow probes were custom-made from Perspex and equipped with 20-MHz Doppler crystals (0.5 mm; Iowa Doppler products, Iowa City, IA) glued to a hole in the cuff. To distend the stomach, an intragastric distensible nitrile balloon of a fixed upper volume was placed in the stomach as previously described for rainbow trout (36). The surgical protocol was slightly modified to further refine the surgery and minimize the effects on the animal. No incision was made in the body wall, and instead the wire guide was inserted through the plastic tubing placed in the mouth. Once inside the stomach, the wire guide was used to pierce through the stomach wall as well as the body wall. The catheter attached to the nitrile balloon was then pulled through the two tissues via the mouth, and the nitrile balloon was then pulled in the stomach. This was done with a minimum of tissue damage, and the integrity was confirmed postmortem.
Experimental protocol.
The fish was allowed to recover for 48 h postsurgery for the cardiovascular parameters to resume baseline values. During the recovery and the experimental protocol, the fish were held in individual opaque chambers (40 dm3) supplied with well-aerated circulating seawater (10°C). The photoperiod was kept at 12:12 h.
One group of short-horn sculpins (n = 10) was, after an initial baseline of 10 min, force-fed filets of Hoki corresponding to a weight of 8% body weight. This is considered to be a normal-sized meal. All cardiovascular parameters were measured continuously for the first 25 h and then at 48, 60, and 72 h.
The other group (n = 8), which had been instrumented with the intragastric balloon, was treated like the fed group except that they received no food. After a baseline of 10 min, they were sham-fed, and as soon as the cardiovascular parameters had recovered the balloon was distended using a volume that corresponded to the weight and volume of the meal given to the feeding group (8–10% body wt). The volume of the balloon was then decreased to mimic the dynamics of the gastric emptying over a 72-h period.
To control for the handling during the feeding, a third group of fish (n = 7) was instrumented as described above. Each individual was removed from the experimental chamber, sham-fed, and then transferred back to the experimental chamber.
Experimental equipment. Pda, central Pven, and gastric pressure were measured using pressure transducers (model DPT-6100; Smiths Medizintechnik, Kirchseeon, Germany) connected to a four-channel amplifier (Somedic, Hörby, Sweden). Calibration was performed against static columns of water. The Doppler flow probes were connected to a directional pulsed Doppler flowmeter (model 545C-4; The University of Iowa). A PowerLab system (model 8S; ADInstruments, Castle Hill, Australia) connected to a personal computer with Chart5 installed (ADInstruments) was used for data acquisition.
Data analysis and statistics. Vascular resistances were calculated from raw data values as Rsys = (Pda – Pven)/CO for the systemic resistance and Ri = (Pda – Pven)/Qcma for splanchnic resistance. Rsys = Ri + Rrest, where Rsys is systemic vascular resistance, Rrest represents the vascular resistance in the system except for the splanchnic resistance (Ri), and Qcma is CMA flow. The following two assumptions were made in these calculations: 1) that the driving force through the intestinal artery is the Pda and 2) that blood viscosity does not change during the experimental protocol. Heart rate (HR) was obtained from pulsating pressure traces, and stroke volume (SV) was calculated as SV = CO/HR. All values shown are mean values ± SE. Blood flows and resistances are presented as relative changes with the baseline set to 100%. The baseline value before distension was averaged and compared with the cardiovascular recording at three different time points (30 min, 15 h, and 60 h, as indicated in Figs. 3–9), using a paired t-test followed by a Holm-Bonferroni test to correct for multiple comparisons. To test for differences between the treatments, a two-sample t-test assuming equal variance was used. Significant difference from the baseline/control was assumed when P < 0.05.
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RESULTS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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Gastric emptying.
When a short-horn sculpin is spontaneously feeding on a meal corresponding to 8–10% of body weight, there is a curvilinear decrease in the stomach content over time (Fig. 2). We see no obvious lag phase at the beginning of feeding as has been reported for other species such as rainbow trout (7, 32). Instead, the decrease seems to be most rapid at the beginning of feeding, and by 5 h only 84.1 ± 3.2% remains. By 24 and 48 h, the amount has declined to 60.9 ± 3.4 and 35.0 ± 1.5%, respectively. Finally, at 72 h after feeding, 13.8 ± 3.2% remains in the stomach. At 72 h, we also estimated the stomach content of the instrumented fish, and most of the ingested food had by then passed into the intestine (20% remaining in stomach).
Influence of feeding. Animals were force-fed a diet of fish meat corresponding to 8–10% body weight, and, to control for the handling, another group (control) was sham-fed before being returned to the experimental chamber.
Within 30 min after feeding, there is a significant increase in the Rcma (24 ± 4%), contributing to an insignificant increase in the gastrointestinal vascular resistance (Rsys; 26 ± 10%) (Figs. 3 and 4). There is a significant increase in CO (28 ± 8%) (Fig. 5) that, together with the increased vascular resistance, leads to an increased Pda that subsides within the next 10 h (Fig. 6). The pressor response after 30 min is 54% from a baseline value of 2.1 ± 0.2 kPa. CO increases at first because of an increase in HR (Fig. 7), and, as HR decreases, the increased CO is maintained through an increase in SV (Fig. 8). Because the increase in the Rcma will only account for part of the increase in the total circulation, the absolute increase is smaller in the gastrointestinal portion, and blood is therefore shunted toward the gastrointestinal circulation, leading to an increase in Qcma at 30 min (27 ± 9%) (Fig. 9). When the resistance of the gastrointestinal vessels declines below baseline, 1 h after feeding, there is a substantial increase in the Qcma. After 15 h, there is a 34 ± 7.0% decrease in Rcma, giving a 63 ± 14% increase in Qcma (Figs. 3 and 9). The increase in Qcma is further corroborated by an increase in CO by 41 ± 11%. This increase is largely dependent on an increase in SV (24 ± 10%), since HR has returned to baseline by 60 h (Figs. 7 and 8). There was no consistent venous response.
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After 30 min, there is a significant increase (P < 0.05) in the Rsys (28 ± 5.7%; Fig. 4). This increase leads to a 19% increase in the Pda, from a baseline of 2.1 ± 0.13 kPa, resembling the response seen in rainbow trout (Fig. 6). In the absence of a simultaneous increase in the Rcma, blood is shunted toward the gastrointestinal tract. The increase in Qcma (27 ± 8%) is, however, not statistically significant (Fig. 9).
After 15 h, there is a significant decrease in CO (37 ± 10%) (Fig. 5) that is due solely to a decrease in SV (33 ± 12%), since there is no change in HR during the experiment (Figs. 7 and 8). The decreased SV was most likely provoked by an increased afterload due to a substantial increase in both the Rsys and the Rcma (123 ± 27 and 82 ± 24%, respectively) at 15 h. As such, the massive increase in vascular resistance differs fundamentally from the decrease in vascular resistance seen in feeding fish. This prohibits an increase in Qcma and also prolongs the pressor response. At between 48 and 72 h, most parameters approach baseline. As with the feeding experiment, the venous response was very inconsistent.
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DISCUSSION |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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-adrenergic increase in both the systemic and the gastrointestinal vascular resistance, leading to a Pda response. This response, which is similar to the initial response seen after feeding in mammals (18, 41), may be of importance in preparing the cardiovascular system for the subsequent hyperemia that occurs as the ingested food is digested and the breakdown products are released in the proximal intestine.
To investigate whether or not this response is also present in another fish species, and one with radically different feeding habits compared with the rainbow trout, we measured key cardiovascular parameters in the short-horn sculpin (M. scorpius), an inactive ambush predator, during both mechanical stomach distension and feeding. When the stomach is distended with a volume corresponding to 8–10% of body weight, there is a rapid increase in both the systemic and gastrointestinal vascular resistance, without a subsequent increase in Q. This increased resistance induces an increased Pda (by 19%) that is both quantitatively and qualitatively similar to the response seen in rainbow trout. In the rainbow trout, this response was -adrenergically mediated (36), given that it could be blocked using the -adrenergic antagonist prasozin. Even though we did not test this hypothesis in the present study, we believe that there should not be any difference, given the similarity of the response.
When the sculpins were fed with fish meat corresponding to the distending volume used, there was an even more substantial dorsal aortic pressor response (54%). The increased pressor response was, in addition to the increasing vascular resistances, further augmented by a simultaneous increase in CO as both SV and HR increased significantly. This agrees with previous results from other inactive ambush predators (3–5) but also more active fish (2) where an increased HR is more important. Eventually, there is a massive decrease in the Rsys that, most likely, is dependent on a large decrease in the gastrointestinal vascular resistance, since the CO continues to increase. This leads to a decrease in the Pda that is not seen with distension of the stomach, where there is instead a massive increase in vascular resistance. The reason for this marked increase in resistance as the distending pressure is removed is as present unknown.
This drop in Rcma coincides well with the entry of food in the proximal intestine as shown by our measurements of gastric emptying (Fig. 2). The subsequent hydrolysis of the food will release amino acids, carbohydrates, and lipids/fatty acids in the intestine. The decrease in vascular resistance and the increased CO facilitate a large increase (92%) in the gastrointestinal blood flow. These results agree well with what has been seen earlier in the red Irish lord (Hemilepidotus hemilepidotus) (5), a related sculpin species. The increase in CO is, however, two times that we report here, and there would most likely be little need for a redistribution of blood in the red Irish lord.
In mammals, it is now commonly accepted that the chemical components of the hydrolyzed food, such as glucose (9, 11, 12, 24, 37), amino acids (24, 37) or fatty acids (11, 24, 37), or a combination of these, are crucial to the postprandial hyperemia as reviewed by Gallavan and Chou (17). Chemical stimuli from food induce a vasodilation in mammals (29) and fish (5), which are local and confined to the position of the food in the gut, although some disparity still exists (1). When chemical stimuli are absent, as during purely mechanical distention of the stomach, we see no decrease in vascular resistance, and hence the blood flow increase does not commence. This observation also agrees with previous mammalian studies (11, 17).
The importance of the mechanical stimulus of ingested food has only been partially investigated in mammals (11, 26–28). It could nevertheless be of importance and stimulate the cardiovascular responses seen during the initial feeding phase (18). These responses could enhance the shunting of blood from less metabolically active tissue such as muscles to the active gastrointestinal tract, as reported by several investigators (16, 41). Because the cardiovascular system is unable to supply all tissues with blood at the same time (14), rearrangements must be made to sustain an increased blood flow to the gut. In fish, this is generally believed to be achieved through an increased CO (3–5). However, the results of the present study and previous results from the rainbow trout (36) indicate that there is also a redistribution of blood from "inactive" organs in addition to the increased CO. Furthermore, this would also partly explain the decreased swimming performance seen in several studies after feeding (4, 15, 38).
The short-horn sculpin is an inactive animal in which the gastrointestinal canal and the surrounding vessels make up a large portion of the animal (Fig. 1). At rest, the vessels of the gut probably receive in excess of the 20–40% of Q seen in several other fish species, such as 40% in Atlantic cod (Gadus morhua) (4), 36% in Chinook salmon (Oncorhyncus tshawytscha) (39), 34% in red Irish lord (H. hemilepidotus) (5), and 30% in sea raven (Hemitripterus americanus) (3) and mammals (21). In contrast, the rainbow trout is an active swimmer, where the gut and the supplying vessels constitute a smaller portion of the animal (40) than in the sculpin. The similarity in the postprandial response between two species with such physiological and phylogenetic differences thus suggests that the response is important, widespread, and probably evolutionarily conserved, especially since similar responses can be seen in mammalian species (18, 42). Most notably, the larger distension of the sculpin stomach leads to a quantitatively similar Rsys and Rcma response compared with the rainbow trout with an increase of 20–30%. The delayed Rsys and Rcma response was not studied in the rainbow trout.
Corrosion casts of the sculpin vascular system (Fig. 1) reveal a separation between the first and second EBA and the third and fourth EBA, connected through a short anastomosis, which could represent a special adaptation the give additional control over the systemic and gastrointestinal blood flow. However, whether this anastomosis is actively controlled or not is presently unknown.
The gastric emptying resembles what has been seen previously (7, 32), and, by 72 h, very little remains in the stomach. The major difference when comparing different studies is the initial phase. We observe no initial lag phase with a period when nothing enters the proximal intestine from the stomach. However, as reported by Bucking and Wood (7), this is most likely because of differences in the water content of the ingested food. Animals that are fed a "wet" diet with a water content of around 70% show very little lag compared with when animals are fed dry pellets (35, 43). Axelsson et al. (2) previously reported that gastric emptying time was delayed in instrumented fish compared with uninstrumented fish, possibly because of stress. In this study, we noted no major difference after 72 h. However, the dynamics could differ, since gastric emptying was only estimated postmortem at 72 h for the instrumented fish. It could, nevertheless, indicate that the animals were not stressed to such an extent that it would affect the postprandial response and gastric emptying.
The fact that we see no lag phase cannot account for the rapid increase in gut blood flow after only 30 min, which occurs well before digested food enters the proximal intestine. This rapid response could indicate the presence of gastric chemoreceptors that sense the existence of food in the stomach. This has not been reported previously in fish, whereas the presence of gastric chemoreceptors in mammals has been confirmed. These are responsible for fine tuning digestion as well as detecting the presence of noxious chemicals/drugs as reviewed (34). Alternatively, it could be that mechanical stimuli are indeed important, so that in the short-horn sculpin the presence of mechanical stimuli alone can stimulate a subsequent increase in the gut blood flow. Such an increase was not seen in the rainbow trout. Nonetheless, there is no significant increase in the gut blood flow when distending the stomach even though there is a trend toward an increase. The experimental protocols used in the previous study (36) on the rainbow trout were also probably too short to rule out this possibility.
Conclusions and Future Perspectives
When mechanically distending the stomach in the short-horn sculpin, a similar pressor response in Pda as was described in the rainbow trout is seen. The pressor response was mediated via an increased vascular resistance in both the systemic and the gut vascular system, where the latter constitutes a substantial portion of the Rsys.
The redistribution of blood in combination with an increased CO would enable the large increase seen in gut blood flow in fish as well as in mammals. However, it is not yet known what chemical stimuli induce this postprandial hyperemia although studies in mammals indicate that there are indeed differences depending on the composition of the diet. Future studies in fish would thus have to focus more on the chemically induced hyperemia and also further on the neural and endocrine regulation of postprandial hyperemia in both mammals and fish.
Ultimately, this will not only lead to a greater knowledge about the blood flow regulation in fish (and mammals), but it will also enable the development of more effective feeds and treatments for different pathological conditions affecting growth.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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REFERENCES |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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  A. Grans, F. Albertsson, M. Axelsson, and C. Olsson Postprandial changes in enteric electrical activity and gut blood flow in rainbow trout (Oncorhynchus mykiss) acclimated to different temperatures J. Exp. Biol., August 15, 2009; 212(16): 2550 - 2557. [Abstract] [Full Text] [PDF]
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 H. Seth, E. Sandblom, and M. Axelsson Nutrient-induced gastrointestinal hyperemia and specific dynamic action in rainbow trout (Oncorhynchus mykiss)--importance of proteins and lipids Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2009; 296(2): R345 - R352. [Abstract] [Full Text] [PDF]
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