Journal of Comparative and Physiological Psychology 1975, Vol. 88, No. 1, 231-238
PRODUCTION OF SATIETY WITH SMALL 1INTRADUODENAL INFUSIONS IN THE RAT CHARLES T. SNOWDON2 University oj Wisconsin Rats equipped with chronic intragastric and intraduodenal catheters received small infusions of various solutions through one catheter during spontaneous meals. Regardless of which compartment or which solution was infused, the animals maintained a constant daily nutrient intake. However, all hypertonic solutions reduced mean meal size and increased the frequency of feeding when injected intraduodenally, while only nutritive solutions reduced mean meal size when infused intragastrically. Water ingestion varied with the effective osmotic pressure of the injected solutions, but there were no differences in water ingestion as a function of the compartment infused. These data suggest both the presence of a duodenal satiety mechanism and the validity of interpreting the meal patterns of vagotomized rats eating a liquid diet as resulting from the rapid emptying of the diet into the duodenum.
Snowdon and Epstein (1970) found a pattern of small frequent meals in vagotomized rats ingesting a liquid diet. Snowdon (1970) subsequently showed that the gastric emptying time with liquid diets in the vagotomized rats was faster than that of nonvagotomized rats. It was argued that this rapid emptying of food into the duodenum activated a nonvagally mediated satiety mechanism that inhibited feeding. Ehman, Albert, and Jamieson (1971) developed a technique for duodenal fistulation and injected 17-hr, food-deprived rats intraduodenally with food, cellulose, hypertonic sodium chloride, hypertonic glucose, and isotonic saline. They found that both nonnutritive bulk and hypertonic solutions infused into the duodenum reduced subsequent food intake, and they argued that bulk and osmotic pressure served to inhibit feeding. Since the suggestive evidence of a duodenal satiety mechanism was found in the vagotomized preparation, the vagus nerve could not be involved as a feedback loop. Therefore, the possibility exists that the re1 Supported by U.S. Public Health Service Grant lease of some duodenal hormone serves to MH-23,197. I am grateful to G. P. Smith, R. S. Wampler, and J. Gibbs for their criticisms, of an signal satiety. Davis, Gallagher, Ladove, earlier draft. These data were presented to the and Turausky (1969), using a transfusion meeting of the Psychonomics Society, St. Louis, technique, found that the blood from rats Missouri, in November 1973. that had been fed to satiety suppressed 2 Requests for reprints should be sent to Charles T. Snowdon, Department of Psychology, Charter feeding when infused into food-deprived at Johnson, University of Wisconsin, Madison, rats, but that blood from food-deprived rats did not initiate feeding in sated animals. Wisconsin 53706. 231
There have been several recent suggestions that the intestines might be involved in the regulation of food intake. Balagura and Fibiger (1968) injected rats with 5, 12, or 25 ml. of water or diet and found that only the 5-ml. injection remained in the stomach immediately after the termination of intubation. Significant volumes of the other injections immediately entered the intestines. Although presenting no demonstration of a feeding inhibition in their own data, they argued that many previous studies claiming to show a satiety effect due to gastric loading may really have been activating intestinal mechanisms. Subsequently, Balagura and Coscina (1969) showed a greater suppression of performance in a feeding situation with 12-ml. intubations of a liquid diet relative to 12 ml. of isotonic saline or to 6 ml. of diet. They argued that the fast emptying of a 12-ml. load into the intestines and the nutritive qualities of the diet might be operating to produce satiety via some intestinal mechanism.
232
CHARLES T. SNOWDON
Thus, their data indicated the presence of a satiety hormone. Schally, Redding, Lucien, and Meyer (1967) showed that injections of purified enterogastrone inhibited feeding in food-deprived mice. However, since the chemical structure of enterogastrone has not been specified and the process of purification is difficult, little subsequent work has been done with this substance. Recently, Gibbs, Young, and Smith (1973a) demonstrated that injections of another duodenal hormone, cholecystokinin, induced a cessation of feeding in deprived rats without any indications that the hormone had aversive properties. Subsequently, Gibbs, Young, and Smith (1973b) have used rats with open gastric fistulas and which, therefore, ingest large quantities of food. These animals immediately suppressed sham feeding when injected with cholecystokinin. Both cholecystokinin and a synthetic C-terminal octapeptide of cholecystokinin operated with equal effectiveness. Finally, they have shown that the suppression of sham feeding after cholecystokinin injections is similar to the cessation induced by intraduodenal injections of diet (Smith, Gibbs, & Young, 1974). The present study presents additional evidence for a duodenal satiety mechanism. Previous studies have used duodenal injections to suppress feeding only in food-deprived rats. In this study solutions were infused into the duodenum coincident with all spontaneous meals in nondeprived rats, both to determine whether a similar suppressive effect would appear during spontaneous feeding, and to see whether the meal patterns obtained with vagotomized rats feeding on a liquid diet could be obtained in normal rats by mimicking rapid stomach emptying. Second, Ehman, et al. (1971) found that some substances (especially glucose) had an effect with duodenal injection that seemed to mimic their effects intragastrically. In the present experiment each series of infusions was also given to the same animal intragastrically so that a direct comparison between gastric and duodenal mechanisms might be made. Finally, substances of varying nutritive and osmotic
effects were used to determine the role of osmotic pressure versus nutritive or bulk factors. METHOD
Subjects The subjects were 9 male rats obtained from the Sprague-Dawley Co., Madison, Wisconsin. Their weights ranged 293-485 gm. before surgery (M = 405.3 gm.).
Procedure Surgery. Chronic nasopharyngeal intragastric catheters were installed in accordance with the procedure described by Epstein (1967). The chronic intraduodenal catheter was constructed as follows: A Va-in. piece of polyethylene (PE) 260 tubing was flanged at each end by holding it over a match flame or soldering iron tip. The heat-softened and expanded tubing was pressed against a flat surface to complete the flange. In the center of this piece a 1-in. square piece of hernia mesh (Usher's Marlex) was cemented with dental cement (Figure 1A). Scar tissue would grow through the mesh providing a solid anchoring of the fistula. A 7-in. length of PE 50' tubing was passed through a 1-in. piece of 17-ga. stainless steel needle tubing bent in a 90° angle. A flange was made in one end of the PE 50 tubing and pulled back over one end of the metal tubing and a 1-in. piece of polyvinyl chloride tubing (!/{$ in. bore, Vie in. wall) was pulled over the flange and glued with epoxy adhesive. Finally, a piece of wire was placed in the lumen of the PE 50 tube and a 90° bend made in the tubing 4% in. from the bend in the metal tubing. The animal was then anesthetized under sodium pentobarbital (Nembutal, 4-mg/100-gm), a scalp incision made, the skull cleared, and 4 screws imbedded in the skull. A second incision was made on the abdomen 1 in. long and l/z in. to the surgeon's left of the midline and starting just below the rib cage. A subcutaneous tunnel was made from this incision to the scalp incision, and the end of the PE 50 tube was pulled through with a hemostat. The PE 50 tubing was cut to a length of 3A in. from the 90° bend and the flanged PE 260 piece slipped over the end. A flange was made in the PE 50 tube large enough so that the PE 260 tube could not slip off, (Figure IB) and finally, the PE 50 and PE 260 tube flanges were fused together with heat. Care was taken that the lumen of the PE 50 tube remained open, and that no openings existed between the PE 260 and PE 50 tubes (Figure 1C). The duodenum was exposed and bathed continuously with isotonio saline. A purse-string suture was constructed on the ventral surface, a slit made in the center of the suture and the flange of the catheter inserted. The suture was drawn and tied
DUODENAL SATIETY MECHANISM and 2 additional sutures made. The hernia mesh remained on the outside of the duodenum. The duodenum was returned to its normal place, the abdominal wall was sutured closed, and the skin incision was closed. The metal tubing part of the catheter was then cemented to the skull with dental cement. Prophylactic doses of .4-ml. penicillin were given following surgery. It was necessary to construct a cap to seal the opening of the duodenal fistula. A Vz-in. length of 15-ga. needle tubing was crimped at one end and inserted into the polyvinyl tube. It was also necessary to flush the duodenal fistula at least every other day, and preferably, daily, with 1-2 ml. of warm water. If this was not done and if the animals were being maintained on a solid diet, the tubing would become permanently clogged. The animals recovered completely from the surgery within 2 wk. and could be tested for upward of 2 mo. Testing. The animals were placed in a feeding chamber similar to that described by Snowdon (1969) in which depression of a lever would deliver a liquid diet through a spout located directly above the lever for as long as the bar was held depressed. Printing counters recorded the duration of each meal (which could be readily converted to meal volume) and the duration of the intervals between meals. A peristaltic pump delivered the diet to the spout at a constant rate of .6 ml/min. This rate is slightly slower than the rat's normal rate of ingestion. To test the effects of infusions of solutions into the stomach or duodenum a second channel of each pump was connected to pass the appropriate test solution from a reservoir through a counter-weighted tube above the animal, through a swivel joint (Epstein & Teitelbaum, 1962) into the opening of either the intragastric or intraduodenal tube. All solutions were at room temperature (23.5°C) as they entered the animal. Solutions were infused only during the period when the animals ingested spontaneously. This has been shown by Quartermain, Kissileff, Shapiro, and Miller (1971) to be the best infusion procedure to obtain precision of caloric compensation for the infusion. After each rat had learned to hold the level depressed to obtain all of its food intake and had showed stable intake for at least 1 week, the infusions were begun. The solutions used were isotonic saline (.9%), .5 M NaCl (2.9%), 1 M glucose (18.0%), 1 M urea (6.0%), and the General Biochemicals (GBI) 116 EC diet that the animals were ingesting orally. The .5 M NaCl, 1 M glucose, and 1 M urea solutions were selected to have theoretically similar osmotic pressures (1 osmole/1), although they have differing physiological osmotic effects. The GBI 116 EC diet had a considerably greater osmotic pressure (approximately 3.0 osmoles/1). The glucose solution contained .69 kcal/ ml and the GBI diet 1.5 kcal/ml. The infusions were carried out for 3 days at a time. A solution would be injected into 1 compartment for 3 days,
233 FLANGED PE E60 TUBE MARLEX MESH
FLANGE PE 260 TUBE MESH
B.
PE 50 TUBE FUSED PE 50
c.
AND PE 260 TUBES
PE 50 TUBE
FIGURE 1. Steps in the construction of an intraduodenal catheter. (A: The flanged PE 260 tubing with hernia mesh cemented at center. B: The PE SO tubing passed through the PE 260 collar and heat flared. C: The PE 50 and PE 260 pieces heat fused together.) then into the other compartment for 3 days. This would be followed by 3 days with no infusions before the next solution would be injected. The order of duodenal or gastric infusions was counterbalanced over subjects within a given solution condition and counterbalanced within a subject over the series of solutions. Readings were taken between 1 p.m. and 3 p.m. each day and the testing room was maintained on a 12-hr.: 12-hr, light/dark cycle over the entire experiment. Since varying numbers of animals completed each solution condition, data were analyzed comparing intragastric and intraduodenal data with the control values for each condition. No comparisons were possible across solution conditions. To evaluate the significance of differences, t tests were used.
RESULTS Feeding Behavior Table 1 presents the data on the mean calories ingested daily in each condition.
CHARLES T. SNOWDON
234
TABLE 1 MEAN DAILY CALORIC INTAKE
Solution
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Biochemicals diet
n
9 4
4 5 6
IntragastricintraIntra- IntraduoduoControl gastric denal infusion infusion denal difference 64.43 64.78 64.78 62.97 64.19
70.97 70.38 64.38 78.71' 73.70
66.58 63.88 55.63 66.00 60.30
ns ns ns .05 ns
* Significantly different from control level, p < .05.
The only difference that appeared was a significant elevation of caloric intake during intragastric glucose infusion relative to both control levels and intraduodenal infusion levels. In no case was there any difference between intraduodenal infusion intake levels and control levels, indicating that animals were capable of regulating their caloric intakes despite duodenal infusions of nutritive substances and that the duodenal infusions did not act aversively to suppress caloric intake. Table 2 presents the data on the mean number of meals per day. Infusions of solutions intragastrically had no effect on the number of meals ingested per day. However, with each of the solutions except the GBI diet there was a significant increase in meal frequency when the solutions were infused intraduodenally. With each of the nonnutritive solutions (isotonic saline, urea, and sodium chloride) there was no significance in the differences between intraduodenal and intragastric infusion values. With glucose infusions the intragastric-intraduodenal difference was significant (p < .02), and with GBI diet the difference barely missed being significant (p > .05). The data for oral meal sizes are presented in Table 3. With intragastric infusions there was no difference from control meal sizes with each of the nonnutritive infusion substances. However, both glucose and GBI infusions significantly suppressed meal size. Regardless of the compartment infused, infusions of nutritive solutions should suppress oral meal sizes if the total daily nu-
trient intake is to remain constant. With intraduodenal infusions all infused solutions produced a significant reduction in meal size, though the suppression of meal size with isotonic saline was considerably less than the suppression with each of the hyperosmotic solutions. Similarly, when oral meal sizes with intragastric infusions were compared with those with intraduodenal infusions, there was a significant difference between the two with urea, glucose, and GBI diet, and a moderately significant difference with isotonic saline. The reduction of meal size found with intragastric infusions of glucose and GBI diets became a significantly greater reduction when the solutions were infused intraduodenally. Thus, the infusion of hyperosmotic substances into the duodenum reduced mean oral meal size and tended to increase meal frequency regardless of the nutrient constituents of the solutions. To a lesser degree isotonic saline infusions into the duodenum also reduced mean oral meal size indicating a secondary effect of bulk.
Water Intake The data on voluntary water ingestion are presented in Figure 2 and Table 4 and are expressed as a mean percentage of control intake levels. The statistical comparisons indicated that there was no difference in water intake as a function of the compartment infused. The effect of a given infused solution on voluntary water ingestion was identical with both intragastric and intraduodenal infusions. Thus, the osTABLE 2 MEAN DAILY NDMBEB OP MEALS
Solution
n
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Biochemicals diet
9 4 4 5 6
IntragastricIntra- IntraintraduoControl gastric denal duoinfusion infusion denal difference 15.83 14.34 14.34 14.40 17.02
17.35 17.00 19.67 15.67 13.80
20.54*« 19.00** 21.67* 18.53* 18.34
* Significantly different from control value, p < .05. ** p < .02.
ns ns ns .02 ns
235
DUODENAL SATIETY MECHANISM
Solution
n
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Biochemicals diet
9 4 4 5 6
IntragastricIntra- IntraintraduoduoControl gastric denal infusion infusion denal difference
2.79 3.01 3.01 2.92 2.64
2.89 2.78 2.32 2.26*** 1.76*
2.32* 2.23*** 1.73*** 1.58*** 1.07***
.05 .01
ns .01 .01
* Significantly different from control, p < .05. * " p < .01.
motic sensitivity of the duodenal satiety mechanism cannot be explained in terms of a differential hydrational response to hyperosmotic stimulation of the duodenum versus the stomach, which might lead to increased gastrointestinal bulk. With isotonic saline and with 1 M glucose there was a significant reduction in voluntary water ingestion, whereas infusion of 1 M NaCl produced a significant elevation of water ingestion. Both 1 M urea and the GBI diet produced water intakes that were not significantly different from noninjection control levels. The failure of GBI infusion to increase water ingestion despite its high osmotic pressure is to be expected since the total volume of orally ingested plus infused diet was no greater than the total orally ingested diet in the control condition. DISCUSSION The infusion of small volumes of osmotically potent substances into the duodenum of rats during the course of spontaneous feeding suppressed the size of meals to a significantly greater degree than the infusion of similar substances intragastrically. None of the nonnutritive osmotic solutions suppressed feeding when infused intragastrically. The suppression of meal sizes with intragastric infusions of nutritive solutions most likely represents a compensation necessary to maintain nutritive constancy rather than indicating a specific nutrient detector in the stomach. The same logic would apply to the greater reduction in meal size found with the intraduodenal infusion
of nutritive substances. The range of differences in mean meal size between intragastric and intraduodenal infusions remained constant across both nutritive and nonnutritive hyperosmotic solutions (.55.70 ml.). Thus, the suppressive effect of intraduodenal injections is primarily due to the hyperosmotic rather than the nutritive qualities of the diet. The finding that rats show caloric compensation for intragastric infusions of glucose and GBI diet during spontaneous meals confirms the results of Quartermain et al. (1971). A similar caloric compensation with nutrient infusion to the duodenum indicates that the same precision of compensation can be obtained with duodenal infusion. The results obtained here with infusions during spontaneous meals over a 3-day period are quite similar to those found by Ehman et al. (1971) who used 3-ml. injections of substances into 17-hr, fooddeprived rats. They found that hypertonic glucose and saline were equally effective in suppressing feeding and, from preliminary results, reported a similar effect with hyperSALINE
UREA
NaC
GLUCOSE
GBI
O 300 at
u. 0
1
Z u 200
un
Ul
a.
INTAKE -
TABLE 3 MEAN ORAL MEAL SIZE (IN MILLILITBES)
d
£100
n
| il IG ID
IG ID
!
1
ID H
1
IB IG ID
IG ID
FIGURE 2. Daily water ingestion as percentage of control levels. (GBI denotes General Biochemicals diet.)
236
CHARLES T. SNOWDON TABLE 4 WATER INTAKE AS PERCENTAGE OF CONTROL CONDITION
Solution
n
gastric infusion
IntraIntragastricduodenal intrainfusion duodenal difference
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Bioehemicals diet
9 4 4 5 6
40.2*** 140.2 275.7** 35.8*** 85.4
49.3** 196.8 398.5* 36.8*** 138.2
Intra-
ns ns ns ns ns
* Significantly different from control, p < .05. •• p < .02. " * p < .01.
tonic urea. A greater degree of suppression with glucose and with the GBI diet in the present experiment can be accounted for in terms of delayed feedback of absorbed nutrient levels operating to reduce the size of successive meals. In addition, Ehman et al. (1971) reported that bulk by itself was sufficient to inhibit feeding. No direct data on bulk are available from the present data, but the small reduction of meal size that resulted from intraduodenal injections of isotonic saline relative to the effects of intragastric injections would indicate that bulk might be a stimulus that inhibits feeding, though not as potent a factor as osmotic pressure. Furthermore, Ehman et al. (1971) found a suppressing effect of bulk when using 3-ml. injections every 15 min. for 2 hr., but found that a single 3-ml. injection of hypertonic solution was sufficient to inhibit feeding. The findings with glucose infusion also agree with those of Yin and Tsai (1973), showing no feeding suppression with intragastric glucose but a clear suppression with intraduodenal infusions of concentrations approaching 1 M. Reduced meal size and increased frequency of feeding with duodenal infusions suggest that the interpretation of the changed meal patterns in vagotomized rats eating a liquid diet in terms of rapid gastric emptying activating a duodenal satiety mechanism (Snowdon, 1970) is a plausible one. Snowdon (1970) also suggested that the activation of such a satiety mechanism might be aversive to the rat. The determina-
tions of Smith ct al. (1974) that cholecystokinin injections produce a "behavioral tranquilization" and that intraduodenal diet injections have produced similar tranquility in the 2 animals they studied, plus the results of the present study (in which total daily food intake was not depressed by duodenal injections) would suggest that duodenal stimulation is not aversive. In addition, Gibbs et al. (1973a) have shown that repeated injections of cholecystokinin do not act to form a conditioned aversion. This does not prove that intraduodenal injections would be nonaversive, but this finding and the maintenance of normal daily caloric intake with intraduodenal infusions would indicate a lack of aversiveness. A gastric osmotic satiety effect has been established in several previous studies. Smith and Duffy (1957), injecting volumes generally between 10 and 15 ml., found that both bulk and hypertonicity inhibited eating. However, in light of the findings of Balagura and Fibiger (1968) that gastric loads of this size enter the duodenum immediately, the Smith and Duffy data can perhaps best be considered as affecting both stomach and intestinal mechanisms. Schwartzbaum and Ward (1958) used only 5-ml. injections and found an increasing suppression of food intake with increasing osmolarity that was independent of the substance used. This effect was determined in 24-hr, food-deprived rats over a 45-min. test period. The equivalence of glucose and sodium chloride as suppressive agents would presumably be due to the lack of nutrient feedback of glucose absorption or metabolism over 45 min. Such feedback would not be equivalent over a 3-day period of ad-lib feeding, as used in the present study. McCleary (1953) reported a suppression of glucose intake following intragastric intubation of 3 ml. of solution with similar concentration effects resulting from glucose, urea, and sodium chloride solutions. In the last 2 studies injection volumes were small enough to minimize any duodenal stimulation. The results of these short-term intragastric loading experiments correspond closely to the effects obtained with infusions occurring during all spontaneous meals in a
DUODENAL SATIETY MECHANISM
3-day period, both intragastrically and intraduodenally in the present experiment. The mechanisms of satiety induction in the duodenum and the stomach appear to be similar, although the mechanism in the duodenum is much more sensitive. The water intake data generally agree with other investigators. Ehman et al. (1971) reported food/water ratios that were similar for both hypertonic glucose and isotonic saline infusions into the duodenum. In the present study, infusion of both these solutions reduced the voluntary water intake of the animals below control levels. Both Ehman et al. and the present experiment found increased water ingestion when hypertonic saline was injected. With urea, Ehman and his co-workers report a preliminary study that gave water intake levels comparable to those of isotonic saline injections; in the present experiment, infusions of urea produced a nonsignificant increase in voluntary water intake. The different findings with respect to urea could be resolved by noting the time differences in the 2 experiments. Ehman et al. observed feeding and drinking over a 2.5-hr, period following a single duodenal infusion, while the present experiments provided an infusion with every meal over 6 days. Since the dipsogenic action of urea develops more slowly than other hyperosmotic stimuli, its effects on drinking might not have appeared by the end of a 2.5-hr, test period. Substances that acted similarly to inhibit feeding when injected intraduodenally had a diverse pattern of effects on water ingestion and extremely small infusions (as little as 1.1 ml. [M] with the GBI diet) inhibited feeding. However, intraduodenal infusions of isotonic saline did produce a small reduction in mean size. These findings both suggest that bulk or volume resulting from injection size or from water drawn into the duodenum from osmotically potent stimuli distending the duodenum is not the major mechanism involved in duodenal satiety. A sensory system responsive to the immediate osmotic effects of solutions would seem a more effective mechanism. Since the satiety effect is also seen clearly following vagotomy, information can be carried to the central nervous system only by the splanch-
237
nic nerves or by a hormone. The work of Gibbs et al. (1973a, 1973b) and Smith et al. (1974) raise cholecystokinin as a likely candidate. A direct relationship between osmotic stimulation of the duodenum and cholecystokinin release now needs to be shown as well as the effect of both duodenal loading and cholecystokinin administration on the central neurological controls of feeding. REFERENCES Balagura, S., & Coseina, D. V. Influence of gastrointestinal loads on meal-eating patterns. Journal oj Comparative and Physiological Psychology, 1969, 69,101-106. Balagura, S., & Fibiger, H. C. Tube feeding: Intestinal factors in gastric loading. Psychonomic Science, 1968, 10, 373-374. Davis, J. D., Gallagher, R. J., Ladove, R. F., & Turausky, A. J. Inhibition of food intake by a humoral factor. Journal oj Comparative and Physiological Psychology, 1969, 67, 407-414. Ehman, G. K., Albert, D. J., & Jamieson, J. L. Injections into the duodenum and the induction of satiety in the rat. Canadian Journal of Psychology, 1971, 25,147-166. Epstein, A. N. Feeding without oropharyngeal sensations. In M. R. Kare & 0. Mailer (Eds.), The chemical senses and nutrition. Baltimore: Johns Hopkins University Press, 1967. Epstein, A. N., & Teitelbaum, P. A water tight swivel joint permitting chronic injections into moving animals. Journal of Applied Physiology, 1962,17, 171-172. Gibbs, J., Young, R. C., & Smith, G. P. Cholecystokinin decreases food intake in rats. Journal of Comparative and Physiological Psychology, 1973, 84, 488-495. (a) Gibbs, J., Young, R. C., & Smith, G. P. Cholecystokinin elicits satiety in the rat with open gastric fistula. Nature, 1Q73, 245, 323-325. (b) McCleary, R. A. Taste and post-ingestion factors in specific-hunger behavior. Journal oj Comparative and Physiological Psychology, 1953, 46, 411-421. Quartermain, D., Kissileff, H. R., Shapiro, R., & Miller, N. E. Suppression of food intake with intragastrio loading: Relation to natural feeding cycle. Science, 1971, 173, 941-943. Schally, A. V., Redding, T. W., Lucien, H. W., & Meyer, J. Enterogastrone inhibits feeding by fasted mice. Science, 1967, 157, 210-211. Schwartzbaum, J. S., & Ward, H. P. An osmotic factor in the regulation of food intake in the rat. Journal oj Comparative and Physiological Psychology, 1958, 51, 555-560. Smith, G. P., Gibbs, J., & Young, R. C. Cholecystokinin and "intestinal satiety" in the rat. Federation Proceedings 1974, 33, 1146-1149. Smith, M., & Duffy, M. Some physiological factors that regulate eating behavior. Journal oj Com-
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parative and Physiological Psychology, 1957, 50, 601-608. Snowdon, C. T. Motivation, regulation, and the control of meal parameters with oral and intragastric feeding. Journal of Comparative and Physiological Psychology, 1969, 69, 91-100. Snowdon, C. T. Gastrointestional sensory and motor control of food intake. Journal oj Comparative and Physiological Psychology, 1970, 71, 6S-76.
Snowdon, C. T., & Epstein, A. N. Oral and intragastric feeding in vagotomized rats. Journal oj Comparative and Physiological Psychology, 1970, 71, 59-67. Yin, T. H., & Tsai, C. T. Effects of glucose on feeding in relation to routes of entry in rats. Journal oj Comparative and Physiological Psychology, 1973, 85, 258-264. (Received November 9, 1973)
PRODUCTION OF SATIETY WITH SMALL 1INTRADUODENAL INFUSIONS IN THE RAT CHARLES T. SNOWDON2 University oj Wisconsin Rats equipped with chronic intragastric and intraduodenal catheters received small infusions of various solutions through one catheter during spontaneous meals. Regardless of which compartment or which solution was infused, the animals maintained a constant daily nutrient intake. However, all hypertonic solutions reduced mean meal size and increased the frequency of feeding when injected intraduodenally, while only nutritive solutions reduced mean meal size when infused intragastrically. Water ingestion varied with the effective osmotic pressure of the injected solutions, but there were no differences in water ingestion as a function of the compartment infused. These data suggest both the presence of a duodenal satiety mechanism and the validity of interpreting the meal patterns of vagotomized rats eating a liquid diet as resulting from the rapid emptying of the diet into the duodenum.
Snowdon and Epstein (1970) found a pattern of small frequent meals in vagotomized rats ingesting a liquid diet. Snowdon (1970) subsequently showed that the gastric emptying time with liquid diets in the vagotomized rats was faster than that of nonvagotomized rats. It was argued that this rapid emptying of food into the duodenum activated a nonvagally mediated satiety mechanism that inhibited feeding. Ehman, Albert, and Jamieson (1971) developed a technique for duodenal fistulation and injected 17-hr, food-deprived rats intraduodenally with food, cellulose, hypertonic sodium chloride, hypertonic glucose, and isotonic saline. They found that both nonnutritive bulk and hypertonic solutions infused into the duodenum reduced subsequent food intake, and they argued that bulk and osmotic pressure served to inhibit feeding. Since the suggestive evidence of a duodenal satiety mechanism was found in the vagotomized preparation, the vagus nerve could not be involved as a feedback loop. Therefore, the possibility exists that the re1 Supported by U.S. Public Health Service Grant lease of some duodenal hormone serves to MH-23,197. I am grateful to G. P. Smith, R. S. Wampler, and J. Gibbs for their criticisms, of an signal satiety. Davis, Gallagher, Ladove, earlier draft. These data were presented to the and Turausky (1969), using a transfusion meeting of the Psychonomics Society, St. Louis, technique, found that the blood from rats Missouri, in November 1973. that had been fed to satiety suppressed 2 Requests for reprints should be sent to Charles T. Snowdon, Department of Psychology, Charter feeding when infused into food-deprived at Johnson, University of Wisconsin, Madison, rats, but that blood from food-deprived rats did not initiate feeding in sated animals. Wisconsin 53706. 231
There have been several recent suggestions that the intestines might be involved in the regulation of food intake. Balagura and Fibiger (1968) injected rats with 5, 12, or 25 ml. of water or diet and found that only the 5-ml. injection remained in the stomach immediately after the termination of intubation. Significant volumes of the other injections immediately entered the intestines. Although presenting no demonstration of a feeding inhibition in their own data, they argued that many previous studies claiming to show a satiety effect due to gastric loading may really have been activating intestinal mechanisms. Subsequently, Balagura and Coscina (1969) showed a greater suppression of performance in a feeding situation with 12-ml. intubations of a liquid diet relative to 12 ml. of isotonic saline or to 6 ml. of diet. They argued that the fast emptying of a 12-ml. load into the intestines and the nutritive qualities of the diet might be operating to produce satiety via some intestinal mechanism.
232
CHARLES T. SNOWDON
Thus, their data indicated the presence of a satiety hormone. Schally, Redding, Lucien, and Meyer (1967) showed that injections of purified enterogastrone inhibited feeding in food-deprived mice. However, since the chemical structure of enterogastrone has not been specified and the process of purification is difficult, little subsequent work has been done with this substance. Recently, Gibbs, Young, and Smith (1973a) demonstrated that injections of another duodenal hormone, cholecystokinin, induced a cessation of feeding in deprived rats without any indications that the hormone had aversive properties. Subsequently, Gibbs, Young, and Smith (1973b) have used rats with open gastric fistulas and which, therefore, ingest large quantities of food. These animals immediately suppressed sham feeding when injected with cholecystokinin. Both cholecystokinin and a synthetic C-terminal octapeptide of cholecystokinin operated with equal effectiveness. Finally, they have shown that the suppression of sham feeding after cholecystokinin injections is similar to the cessation induced by intraduodenal injections of diet (Smith, Gibbs, & Young, 1974). The present study presents additional evidence for a duodenal satiety mechanism. Previous studies have used duodenal injections to suppress feeding only in food-deprived rats. In this study solutions were infused into the duodenum coincident with all spontaneous meals in nondeprived rats, both to determine whether a similar suppressive effect would appear during spontaneous feeding, and to see whether the meal patterns obtained with vagotomized rats feeding on a liquid diet could be obtained in normal rats by mimicking rapid stomach emptying. Second, Ehman, et al. (1971) found that some substances (especially glucose) had an effect with duodenal injection that seemed to mimic their effects intragastrically. In the present experiment each series of infusions was also given to the same animal intragastrically so that a direct comparison between gastric and duodenal mechanisms might be made. Finally, substances of varying nutritive and osmotic
effects were used to determine the role of osmotic pressure versus nutritive or bulk factors. METHOD
Subjects The subjects were 9 male rats obtained from the Sprague-Dawley Co., Madison, Wisconsin. Their weights ranged 293-485 gm. before surgery (M = 405.3 gm.).
Procedure Surgery. Chronic nasopharyngeal intragastric catheters were installed in accordance with the procedure described by Epstein (1967). The chronic intraduodenal catheter was constructed as follows: A Va-in. piece of polyethylene (PE) 260 tubing was flanged at each end by holding it over a match flame or soldering iron tip. The heat-softened and expanded tubing was pressed against a flat surface to complete the flange. In the center of this piece a 1-in. square piece of hernia mesh (Usher's Marlex) was cemented with dental cement (Figure 1A). Scar tissue would grow through the mesh providing a solid anchoring of the fistula. A 7-in. length of PE 50' tubing was passed through a 1-in. piece of 17-ga. stainless steel needle tubing bent in a 90° angle. A flange was made in one end of the PE 50 tubing and pulled back over one end of the metal tubing and a 1-in. piece of polyvinyl chloride tubing (!/{$ in. bore, Vie in. wall) was pulled over the flange and glued with epoxy adhesive. Finally, a piece of wire was placed in the lumen of the PE 50 tube and a 90° bend made in the tubing 4% in. from the bend in the metal tubing. The animal was then anesthetized under sodium pentobarbital (Nembutal, 4-mg/100-gm), a scalp incision made, the skull cleared, and 4 screws imbedded in the skull. A second incision was made on the abdomen 1 in. long and l/z in. to the surgeon's left of the midline and starting just below the rib cage. A subcutaneous tunnel was made from this incision to the scalp incision, and the end of the PE 50 tube was pulled through with a hemostat. The PE 50 tubing was cut to a length of 3A in. from the 90° bend and the flanged PE 260 piece slipped over the end. A flange was made in the PE 50 tube large enough so that the PE 260 tube could not slip off, (Figure IB) and finally, the PE 50 and PE 260 tube flanges were fused together with heat. Care was taken that the lumen of the PE 50 tube remained open, and that no openings existed between the PE 260 and PE 50 tubes (Figure 1C). The duodenum was exposed and bathed continuously with isotonio saline. A purse-string suture was constructed on the ventral surface, a slit made in the center of the suture and the flange of the catheter inserted. The suture was drawn and tied
DUODENAL SATIETY MECHANISM and 2 additional sutures made. The hernia mesh remained on the outside of the duodenum. The duodenum was returned to its normal place, the abdominal wall was sutured closed, and the skin incision was closed. The metal tubing part of the catheter was then cemented to the skull with dental cement. Prophylactic doses of .4-ml. penicillin were given following surgery. It was necessary to construct a cap to seal the opening of the duodenal fistula. A Vz-in. length of 15-ga. needle tubing was crimped at one end and inserted into the polyvinyl tube. It was also necessary to flush the duodenal fistula at least every other day, and preferably, daily, with 1-2 ml. of warm water. If this was not done and if the animals were being maintained on a solid diet, the tubing would become permanently clogged. The animals recovered completely from the surgery within 2 wk. and could be tested for upward of 2 mo. Testing. The animals were placed in a feeding chamber similar to that described by Snowdon (1969) in which depression of a lever would deliver a liquid diet through a spout located directly above the lever for as long as the bar was held depressed. Printing counters recorded the duration of each meal (which could be readily converted to meal volume) and the duration of the intervals between meals. A peristaltic pump delivered the diet to the spout at a constant rate of .6 ml/min. This rate is slightly slower than the rat's normal rate of ingestion. To test the effects of infusions of solutions into the stomach or duodenum a second channel of each pump was connected to pass the appropriate test solution from a reservoir through a counter-weighted tube above the animal, through a swivel joint (Epstein & Teitelbaum, 1962) into the opening of either the intragastric or intraduodenal tube. All solutions were at room temperature (23.5°C) as they entered the animal. Solutions were infused only during the period when the animals ingested spontaneously. This has been shown by Quartermain, Kissileff, Shapiro, and Miller (1971) to be the best infusion procedure to obtain precision of caloric compensation for the infusion. After each rat had learned to hold the level depressed to obtain all of its food intake and had showed stable intake for at least 1 week, the infusions were begun. The solutions used were isotonic saline (.9%), .5 M NaCl (2.9%), 1 M glucose (18.0%), 1 M urea (6.0%), and the General Biochemicals (GBI) 116 EC diet that the animals were ingesting orally. The .5 M NaCl, 1 M glucose, and 1 M urea solutions were selected to have theoretically similar osmotic pressures (1 osmole/1), although they have differing physiological osmotic effects. The GBI 116 EC diet had a considerably greater osmotic pressure (approximately 3.0 osmoles/1). The glucose solution contained .69 kcal/ ml and the GBI diet 1.5 kcal/ml. The infusions were carried out for 3 days at a time. A solution would be injected into 1 compartment for 3 days,
233 FLANGED PE E60 TUBE MARLEX MESH
FLANGE PE 260 TUBE MESH
B.
PE 50 TUBE FUSED PE 50
c.
AND PE 260 TUBES
PE 50 TUBE
FIGURE 1. Steps in the construction of an intraduodenal catheter. (A: The flanged PE 260 tubing with hernia mesh cemented at center. B: The PE SO tubing passed through the PE 260 collar and heat flared. C: The PE 50 and PE 260 pieces heat fused together.) then into the other compartment for 3 days. This would be followed by 3 days with no infusions before the next solution would be injected. The order of duodenal or gastric infusions was counterbalanced over subjects within a given solution condition and counterbalanced within a subject over the series of solutions. Readings were taken between 1 p.m. and 3 p.m. each day and the testing room was maintained on a 12-hr.: 12-hr, light/dark cycle over the entire experiment. Since varying numbers of animals completed each solution condition, data were analyzed comparing intragastric and intraduodenal data with the control values for each condition. No comparisons were possible across solution conditions. To evaluate the significance of differences, t tests were used.
RESULTS Feeding Behavior Table 1 presents the data on the mean calories ingested daily in each condition.
CHARLES T. SNOWDON
234
TABLE 1 MEAN DAILY CALORIC INTAKE
Solution
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Biochemicals diet
n
9 4
4 5 6
IntragastricintraIntra- IntraduoduoControl gastric denal infusion infusion denal difference 64.43 64.78 64.78 62.97 64.19
70.97 70.38 64.38 78.71' 73.70
66.58 63.88 55.63 66.00 60.30
ns ns ns .05 ns
* Significantly different from control level, p < .05.
The only difference that appeared was a significant elevation of caloric intake during intragastric glucose infusion relative to both control levels and intraduodenal infusion levels. In no case was there any difference between intraduodenal infusion intake levels and control levels, indicating that animals were capable of regulating their caloric intakes despite duodenal infusions of nutritive substances and that the duodenal infusions did not act aversively to suppress caloric intake. Table 2 presents the data on the mean number of meals per day. Infusions of solutions intragastrically had no effect on the number of meals ingested per day. However, with each of the solutions except the GBI diet there was a significant increase in meal frequency when the solutions were infused intraduodenally. With each of the nonnutritive solutions (isotonic saline, urea, and sodium chloride) there was no significance in the differences between intraduodenal and intragastric infusion values. With glucose infusions the intragastric-intraduodenal difference was significant (p < .02), and with GBI diet the difference barely missed being significant (p > .05). The data for oral meal sizes are presented in Table 3. With intragastric infusions there was no difference from control meal sizes with each of the nonnutritive infusion substances. However, both glucose and GBI infusions significantly suppressed meal size. Regardless of the compartment infused, infusions of nutritive solutions should suppress oral meal sizes if the total daily nu-
trient intake is to remain constant. With intraduodenal infusions all infused solutions produced a significant reduction in meal size, though the suppression of meal size with isotonic saline was considerably less than the suppression with each of the hyperosmotic solutions. Similarly, when oral meal sizes with intragastric infusions were compared with those with intraduodenal infusions, there was a significant difference between the two with urea, glucose, and GBI diet, and a moderately significant difference with isotonic saline. The reduction of meal size found with intragastric infusions of glucose and GBI diets became a significantly greater reduction when the solutions were infused intraduodenally. Thus, the infusion of hyperosmotic substances into the duodenum reduced mean oral meal size and tended to increase meal frequency regardless of the nutrient constituents of the solutions. To a lesser degree isotonic saline infusions into the duodenum also reduced mean oral meal size indicating a secondary effect of bulk.
Water Intake The data on voluntary water ingestion are presented in Figure 2 and Table 4 and are expressed as a mean percentage of control intake levels. The statistical comparisons indicated that there was no difference in water intake as a function of the compartment infused. The effect of a given infused solution on voluntary water ingestion was identical with both intragastric and intraduodenal infusions. Thus, the osTABLE 2 MEAN DAILY NDMBEB OP MEALS
Solution
n
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Biochemicals diet
9 4 4 5 6
IntragastricIntra- IntraintraduoControl gastric denal duoinfusion infusion denal difference 15.83 14.34 14.34 14.40 17.02
17.35 17.00 19.67 15.67 13.80
20.54*« 19.00** 21.67* 18.53* 18.34
* Significantly different from control value, p < .05. ** p < .02.
ns ns ns .02 ns
235
DUODENAL SATIETY MECHANISM
Solution
n
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Biochemicals diet
9 4 4 5 6
IntragastricIntra- IntraintraduoduoControl gastric denal infusion infusion denal difference
2.79 3.01 3.01 2.92 2.64
2.89 2.78 2.32 2.26*** 1.76*
2.32* 2.23*** 1.73*** 1.58*** 1.07***
.05 .01
ns .01 .01
* Significantly different from control, p < .05. * " p < .01.
motic sensitivity of the duodenal satiety mechanism cannot be explained in terms of a differential hydrational response to hyperosmotic stimulation of the duodenum versus the stomach, which might lead to increased gastrointestinal bulk. With isotonic saline and with 1 M glucose there was a significant reduction in voluntary water ingestion, whereas infusion of 1 M NaCl produced a significant elevation of water ingestion. Both 1 M urea and the GBI diet produced water intakes that were not significantly different from noninjection control levels. The failure of GBI infusion to increase water ingestion despite its high osmotic pressure is to be expected since the total volume of orally ingested plus infused diet was no greater than the total orally ingested diet in the control condition. DISCUSSION The infusion of small volumes of osmotically potent substances into the duodenum of rats during the course of spontaneous feeding suppressed the size of meals to a significantly greater degree than the infusion of similar substances intragastrically. None of the nonnutritive osmotic solutions suppressed feeding when infused intragastrically. The suppression of meal sizes with intragastric infusions of nutritive solutions most likely represents a compensation necessary to maintain nutritive constancy rather than indicating a specific nutrient detector in the stomach. The same logic would apply to the greater reduction in meal size found with the intraduodenal infusion
of nutritive substances. The range of differences in mean meal size between intragastric and intraduodenal infusions remained constant across both nutritive and nonnutritive hyperosmotic solutions (.55.70 ml.). Thus, the suppressive effect of intraduodenal injections is primarily due to the hyperosmotic rather than the nutritive qualities of the diet. The finding that rats show caloric compensation for intragastric infusions of glucose and GBI diet during spontaneous meals confirms the results of Quartermain et al. (1971). A similar caloric compensation with nutrient infusion to the duodenum indicates that the same precision of compensation can be obtained with duodenal infusion. The results obtained here with infusions during spontaneous meals over a 3-day period are quite similar to those found by Ehman et al. (1971) who used 3-ml. injections of substances into 17-hr, fooddeprived rats. They found that hypertonic glucose and saline were equally effective in suppressing feeding and, from preliminary results, reported a similar effect with hyperSALINE
UREA
NaC
GLUCOSE
GBI
O 300 at
u. 0
1
Z u 200
un
Ul
a.
INTAKE -
TABLE 3 MEAN ORAL MEAL SIZE (IN MILLILITBES)
d
£100
n
| il IG ID
IG ID
!
1
ID H
1
IB IG ID
IG ID
FIGURE 2. Daily water ingestion as percentage of control levels. (GBI denotes General Biochemicals diet.)
236
CHARLES T. SNOWDON TABLE 4 WATER INTAKE AS PERCENTAGE OF CONTROL CONDITION
Solution
n
gastric infusion
IntraIntragastricduodenal intrainfusion duodenal difference
Isotonic saline 1 M urea .5 M NaCl 1 M glucose General Bioehemicals diet
9 4 4 5 6
40.2*** 140.2 275.7** 35.8*** 85.4
49.3** 196.8 398.5* 36.8*** 138.2
Intra-
ns ns ns ns ns
* Significantly different from control, p < .05. •• p < .02. " * p < .01.
tonic urea. A greater degree of suppression with glucose and with the GBI diet in the present experiment can be accounted for in terms of delayed feedback of absorbed nutrient levels operating to reduce the size of successive meals. In addition, Ehman et al. (1971) reported that bulk by itself was sufficient to inhibit feeding. No direct data on bulk are available from the present data, but the small reduction of meal size that resulted from intraduodenal injections of isotonic saline relative to the effects of intragastric injections would indicate that bulk might be a stimulus that inhibits feeding, though not as potent a factor as osmotic pressure. Furthermore, Ehman et al. (1971) found a suppressing effect of bulk when using 3-ml. injections every 15 min. for 2 hr., but found that a single 3-ml. injection of hypertonic solution was sufficient to inhibit feeding. The findings with glucose infusion also agree with those of Yin and Tsai (1973), showing no feeding suppression with intragastric glucose but a clear suppression with intraduodenal infusions of concentrations approaching 1 M. Reduced meal size and increased frequency of feeding with duodenal infusions suggest that the interpretation of the changed meal patterns in vagotomized rats eating a liquid diet in terms of rapid gastric emptying activating a duodenal satiety mechanism (Snowdon, 1970) is a plausible one. Snowdon (1970) also suggested that the activation of such a satiety mechanism might be aversive to the rat. The determina-
tions of Smith ct al. (1974) that cholecystokinin injections produce a "behavioral tranquilization" and that intraduodenal diet injections have produced similar tranquility in the 2 animals they studied, plus the results of the present study (in which total daily food intake was not depressed by duodenal injections) would suggest that duodenal stimulation is not aversive. In addition, Gibbs et al. (1973a) have shown that repeated injections of cholecystokinin do not act to form a conditioned aversion. This does not prove that intraduodenal injections would be nonaversive, but this finding and the maintenance of normal daily caloric intake with intraduodenal infusions would indicate a lack of aversiveness. A gastric osmotic satiety effect has been established in several previous studies. Smith and Duffy (1957), injecting volumes generally between 10 and 15 ml., found that both bulk and hypertonicity inhibited eating. However, in light of the findings of Balagura and Fibiger (1968) that gastric loads of this size enter the duodenum immediately, the Smith and Duffy data can perhaps best be considered as affecting both stomach and intestinal mechanisms. Schwartzbaum and Ward (1958) used only 5-ml. injections and found an increasing suppression of food intake with increasing osmolarity that was independent of the substance used. This effect was determined in 24-hr, food-deprived rats over a 45-min. test period. The equivalence of glucose and sodium chloride as suppressive agents would presumably be due to the lack of nutrient feedback of glucose absorption or metabolism over 45 min. Such feedback would not be equivalent over a 3-day period of ad-lib feeding, as used in the present study. McCleary (1953) reported a suppression of glucose intake following intragastric intubation of 3 ml. of solution with similar concentration effects resulting from glucose, urea, and sodium chloride solutions. In the last 2 studies injection volumes were small enough to minimize any duodenal stimulation. The results of these short-term intragastric loading experiments correspond closely to the effects obtained with infusions occurring during all spontaneous meals in a
DUODENAL SATIETY MECHANISM
3-day period, both intragastrically and intraduodenally in the present experiment. The mechanisms of satiety induction in the duodenum and the stomach appear to be similar, although the mechanism in the duodenum is much more sensitive. The water intake data generally agree with other investigators. Ehman et al. (1971) reported food/water ratios that were similar for both hypertonic glucose and isotonic saline infusions into the duodenum. In the present study, infusion of both these solutions reduced the voluntary water intake of the animals below control levels. Both Ehman et al. and the present experiment found increased water ingestion when hypertonic saline was injected. With urea, Ehman and his co-workers report a preliminary study that gave water intake levels comparable to those of isotonic saline injections; in the present experiment, infusions of urea produced a nonsignificant increase in voluntary water intake. The different findings with respect to urea could be resolved by noting the time differences in the 2 experiments. Ehman et al. observed feeding and drinking over a 2.5-hr, period following a single duodenal infusion, while the present experiments provided an infusion with every meal over 6 days. Since the dipsogenic action of urea develops more slowly than other hyperosmotic stimuli, its effects on drinking might not have appeared by the end of a 2.5-hr, test period. Substances that acted similarly to inhibit feeding when injected intraduodenally had a diverse pattern of effects on water ingestion and extremely small infusions (as little as 1.1 ml. [M] with the GBI diet) inhibited feeding. However, intraduodenal infusions of isotonic saline did produce a small reduction in mean size. These findings both suggest that bulk or volume resulting from injection size or from water drawn into the duodenum from osmotically potent stimuli distending the duodenum is not the major mechanism involved in duodenal satiety. A sensory system responsive to the immediate osmotic effects of solutions would seem a more effective mechanism. Since the satiety effect is also seen clearly following vagotomy, information can be carried to the central nervous system only by the splanch-
237
nic nerves or by a hormone. The work of Gibbs et al. (1973a, 1973b) and Smith et al. (1974) raise cholecystokinin as a likely candidate. A direct relationship between osmotic stimulation of the duodenum and cholecystokinin release now needs to be shown as well as the effect of both duodenal loading and cholecystokinin administration on the central neurological controls of feeding. REFERENCES Balagura, S., & Coseina, D. V. Influence of gastrointestinal loads on meal-eating patterns. Journal oj Comparative and Physiological Psychology, 1969, 69,101-106. Balagura, S., & Fibiger, H. C. Tube feeding: Intestinal factors in gastric loading. Psychonomic Science, 1968, 10, 373-374. Davis, J. D., Gallagher, R. J., Ladove, R. F., & Turausky, A. J. Inhibition of food intake by a humoral factor. Journal oj Comparative and Physiological Psychology, 1969, 67, 407-414. Ehman, G. K., Albert, D. J., & Jamieson, J. L. Injections into the duodenum and the induction of satiety in the rat. Canadian Journal of Psychology, 1971, 25,147-166. Epstein, A. N. Feeding without oropharyngeal sensations. In M. R. Kare & 0. Mailer (Eds.), The chemical senses and nutrition. Baltimore: Johns Hopkins University Press, 1967. Epstein, A. N., & Teitelbaum, P. A water tight swivel joint permitting chronic injections into moving animals. Journal of Applied Physiology, 1962,17, 171-172. Gibbs, J., Young, R. C., & Smith, G. P. Cholecystokinin decreases food intake in rats. Journal of Comparative and Physiological Psychology, 1973, 84, 488-495. (a) Gibbs, J., Young, R. C., & Smith, G. P. Cholecystokinin elicits satiety in the rat with open gastric fistula. Nature, 1Q73, 245, 323-325. (b) McCleary, R. A. Taste and post-ingestion factors in specific-hunger behavior. Journal oj Comparative and Physiological Psychology, 1953, 46, 411-421. Quartermain, D., Kissileff, H. R., Shapiro, R., & Miller, N. E. Suppression of food intake with intragastrio loading: Relation to natural feeding cycle. Science, 1971, 173, 941-943. Schally, A. V., Redding, T. W., Lucien, H. W., & Meyer, J. Enterogastrone inhibits feeding by fasted mice. Science, 1967, 157, 210-211. Schwartzbaum, J. S., & Ward, H. P. An osmotic factor in the regulation of food intake in the rat. Journal oj Comparative and Physiological Psychology, 1958, 51, 555-560. Smith, G. P., Gibbs, J., & Young, R. C. Cholecystokinin and "intestinal satiety" in the rat. Federation Proceedings 1974, 33, 1146-1149. Smith, M., & Duffy, M. Some physiological factors that regulate eating behavior. Journal oj Com-
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parative and Physiological Psychology, 1957, 50, 601-608. Snowdon, C. T. Motivation, regulation, and the control of meal parameters with oral and intragastric feeding. Journal of Comparative and Physiological Psychology, 1969, 69, 91-100. Snowdon, C. T. Gastrointestional sensory and motor control of food intake. Journal oj Comparative and Physiological Psychology, 1970, 71, 6S-76.
Snowdon, C. T., & Epstein, A. N. Oral and intragastric feeding in vagotomized rats. Journal oj Comparative and Physiological Psychology, 1970, 71, 59-67. Yin, T. H., & Tsai, C. T. Effects of glucose on feeding in relation to routes of entry in rats. Journal oj Comparative and Physiological Psychology, 1973, 85, 258-264. (Received November 9, 1973)
