Physiology & Behavior, Vol. 16, pp. 561--568. Pergamon Press and Brain Research Publ., 1976. Printed in the U.S.A.
Gastric Modulation of Gustatory Afferent Activity' JOHN F. GLENN 2 AND ROBERT P. ERICKSON a
Duke University (Received 5 June 1975)
GLENN, J.F. AND R.P. ERICKSON. Gastric modulation of gustatory afferent activity. PHYSIOL. BEHAV. 16(5) 561-568, 1976. - The neural activity evoked by stimulation of the rat's tongue with NaCl, sucrose, HCI, and QHC1 was recorded from 25 small populations of second-order gustatory neurons in the nucleus of the solitary tract, both before and after gastric loading. A slowly developing (several minutes) load-induced depression in the response evoked by the taste solutions was noted, with no significant differences seen between loading with air or 0.3 M NaCl. Differences were evident in the magnitude of the decrement for the various stimuli; the order of decrement was sucrose > NaCl > HC1;the response to QHCI showed no clear decrement. The effect of gastric loading, as tested with air loads, is reversible. The magnitude of the load effect was seen to be less with greater levels of food deprivation. The results of this study therefore, demonstrate the existence of gastric effects on the reactivity of second-order gustatory afferents to chemical stimulation of the rat's tongue. Taste Gastric distention Multineuronal activity
Nucleus of the solitary tract
Food intake
Afferent interaction
factors, determine the organism's feeding response to a foodstuff [321. Given that sensory information from the oral cavity influences the acceptance and rejection of foodstuffs, the question remains - what are the mechanisms by which a given oral stimulus may result in different behaviors? It is possible that, under the influence of both humoral and neural factors, the central integrative, perhaps hypothalamic, mechanisms may simply send a stop or go signal, or graded amounts of stop and go signals, to the mot or centers in the medulla subserving ingestive behavior. However, it is also possible that the hypothalamus and other CNS structures may exert some effect on the afferent input instead of, or in addition to, a more direct effect on the efferent m o t o r system. While such central influences might act on sensory receptors or relays to bias or modify at some level the oropharyngeal sensory input to the feeding reflex, it is also possible that these central influences may be augmented or supplanted by more direct influences exerted on the oral sensory input by ascending visceral afferents from the gut. The juxtaposition of the terminals of afferents for vagally mediated interoceptive sensory information and sensory information from the oropharyngeal cavity in the nucleus tractus solitarius of the hindbrain strongly suggests such direct influences. There are several experiments which demonstrate that
ALTHOUGH feeding behavior historically has been studied from several perspectives, the emphasis of modern research originated within the framework of homeostatic regulatory mechanisms. In particular, the thoughts of Claude Bernard on the constancy of the internal environment have pointed the directions for research on the bases of homeostatic regulation. The rhythmic fluctuations seen in meal frequency, meal size, and the foodstuffs selected consequently are seen as behavioral compensations for deviations from certain criteria of normality in the milieu interieur. While internal, postingestional (PI) signals of b o t h gastric and humoral origin are generally accepted as providing interoceptive information about the intraorganic state, relatively little systematic consideration has been given to the role of external, oropharyngeal factors, especially taste and olfaction, within the mechanisms involved in the regulation of food intake. Yet, it is obvious that the mechanisms controlling ingestive behavior are as inextricably bound to external sensory factors as to internal factors. As examples, the recovery of feeding behavior in brain-damaged animals has been shown to be tastedependent (hypothalamic hyperphagics and aphagics; see [28] ), and satiety has been shown to be sensory specific [5, 14, 15]. Thus, there are both internal (PI) and external (oral and olfactory) classes of situational input which, in conjunction with innate predispositions and experiential
1This paper is based on a doctoral dissertation submitted at Duke University by the first author. This work was supported in part by NSF Grant GB 33464 and in part by NIMH Predoctoral Fellowship 1S01-MH-58159-01. 2j. F. Glenn is presently serving as a Captain in the U.S. Army Medical Service Corps and is assigned to the Behavioral Research Directorate of the U.S. Army Human Engineering Laboratory at Aberdeen Proving Ground, MD 21005. 3Send reprint requests to R. P. Erickson, Professor of Psychology and Associate Professor of Physiology at Duke University, Durham, NC 27706. 561
562
GLENN AND ERICKSON
postingestionally initiated control of the afferent message exists in the gustatory system. Behavioral evidence of postingestional changes in gustatory reactivity has been gathered in both humans [33 ] and dogs [5 ]. Electrophysiological evidence for changes in chemically evoked activity after stomach distention in toads and frogs has been demonstrated in the first-order gustatory afferents of the glossopharyngeal nerve [2,10] and in the gustatory receptor potential [24]. While the mechanisms underlying these behavioral and electrophysiological effects are not clear, it seems obvious that the gustatory message may be altered in some manner by changes in the milieu interieur. Although there is behavioral evidence from humans and dogs for modulation of gustatory afferent information, there has been no electrophysiological verification of intragastrically induced neural changes in the taste system of mammals. Since a large portion of the literature on feeding behavior involves work with rats, it was thought that a demonstration of sensory modulation in that animal would be useful. The nucleus solitarius of the medulla, with its multiple special and general visceral afferent input, would seem to provide a logical locus for integration of visceral and oral information. It is the purpose of this study, therefore, to investigate the effect of intragastric loading on the chemically evoked activity of second-order neurons of the nucleus tractus solitarius of the rat. METHOD
Animals Fifteen female Sprague-Dawley albino rats (Charles River), ranging in weight from 185 to 245 g, were used. Thirteen of the animals were isolated before surgery without access to food, but with ad lib water, for varying periods of time. The two animals not isolated were taken directly from the colony cage, where they had been maintained on an ad lib schedule of both food and water intake. Surgery. The animals were prepared for surgery under a combination of anesthetic (inhalation and barbituate) and neuroleptanalgesic (narcotic analgesic and major tranquilizer) techniques. The animals were first lightly sedated with Fluothane (halothane, Ayerst). Upon sedation, atropine sulfate was given (0.3 mg/kg IP). A single injection of Brevital Sodium (sodium methohexital-Lilly), a rapidly effective, ultra-shortacting barbituate, was used to induce anesthesia (25 mg/kg IP). This dosage provided a surgical level of anesthesia for about 15 min. The trachea was cannulated with polyethlene tubing, and a size 8 Bardic infant intra-gastric feeding tube (C. R. Bard Inc.) was inserted into the esophagus until the tip of the tube passed into the stomach. The animal was then mounted into a head holder which allowed free access to the oral cavity [8]. Innovar Injection (McNeil: narcotic analgesic-fentanyl; neuroleptic-droperidol) was administered (0.1 mg/kg IM) and Anucaine (Calvin Chemical), a long-lasting local anesthetic, was injected in small amounts into the cut edges of the wound to provide a surgical level of anesthesia. The surgical techniques for the exposure of the medulla were carried out under a Zeiss operating microscope by first removing portions of the occipital skull, and then aspirating the cerebellum to expose the floor of the fourth ventricle. The EKG and body temperature were monitored throughout the procedure, and body temperature was maintained between 35 ° and 37.5°C by means of an
electric body warmer. The animal was immobilized with Flaxedil (gallamine triethiodide, Davis and Geck; 60 mg/kg IP) and was artificially respirated by means of a Palmer or Harvard respirator pump. Pneumothorax was performed when necessary to reduce brain movement.
Procedure Recording. The chemically evoked responses in small populations of second-order neurons of the Nucleus Tractus Solitarius were recorded with tungsten microelectrodes (Transidyne General 404-20). The identification of secondorder gustatory activity was made according to the methods and criteria of Doetsch and Erickson [6]. The active electrode was positioned with a three-coordinate micromanipulator, the vertical movements of which were hydraulically driven. A bare nichrome wire positioned on the floor of the fourth ventricle with a second micromanipulator acted as the second recording electrode. Leads from the two electrodes, as well as the animal ground lead, were connected to a Grass P-15 preamplifier. The preamplifier output was fed to a Grass RM-122 low-level preamplifier, the output of which was connected in parallel to an audio monitor, a Tektronix 502 dual-beam oscilloscope, and a magnetic tape recorder (Sony 650-4 or Teac 2340). Stomach loading. The intragastric feeding tube, which was inserted into the cardiac portion of the stomach, was connected to a 10 ml syringe. In order to allow introduction of both chemical and inert loads in the same preparation, simple distention of the stomach was accomplished by the introduction of a volume of air, rather than the more widely used procedure of an intragastric balloon. The peculiarity of the anatomy of the rat's cardiac sphincter, which prevents emesis, probably contributed to the tightness of seal between the intragastric tube and sphincter, and allowed retention of the inert air load throughout the course of the load condition. Surgical exposure of the stomach was made in several rats in a preliminary investigation, and visual inspection showed that this procedure of air loading was reliable and repeatable in that air did not leak out, but could be withdrawn simply and easily by pulling back on the plunger of the syringe. In the volumes injected, the area of distention in these animals seemed to be well confined to the stomach, with no noticeable duodenal distention. The volume injected in the air load condition was approximately 8 ml (range 6 - 1 0 ml). The solution chosen for chemical loading was 0.3 M NaC1. This concentration was deemed appropriate in that it was sufficiently concentrated to remain largely in the stomach for several minutes with reduced clearance relative to water [ 12] without being pathological. The amounts of NaC1 intubed were slightly less than those of air (range 5 . 5 - 8 . 5 ml). Visual inspection of the medulla showed no noticeable movement of the medulla relative to the recording electrode during distention, nor did the neural activity at this time suggest movement. Data collection. Chemical solutions of 0.1 M NaC1, 0.03 M HC1, 0.01 M QHC1, and 1.0M Sucrose were prepared in deionized water, and were introduced by means of a gravity flow system into a chamber fitted around the anterior portion of the tongue. A stimulus presentation consisted of a flow of solution over the tongue for approximately 1 0 - 1 5 sec. Each presentation was followed
GASTRIC MODULATION OF GUSTATORY A F F E R E N T ACTIVITY by a 4 5 - 6 0 sec continuous rinse with deionized water, except in Experiment 1-1 where rinse time was approximately 20 sec. In 14 cases, presentations were made before, during and after removal of the stomach load as a check for the reversibility of distention effects. Data analysis. The records of neural activity consisted of populations of unit responses in which one or several single units were resolvable. The data were thus amenable to analysis by means of a spike counter with a variable threshold amplitude discriminator circuit; the output of the amplitude discriminator was in the form of square wave pulses. For further analysis, the output of the spike counter in each 500 msec period was printed on paper tape by a Franklin Electronics counter. Three measures of activity for each presentation of a stimulus were calculated from the printed spike counts: (1) the average rate of resting (spontaneous) activity during the 3 sec preceding each presentation (B); (2) the number of spikes in the first 3 sec of chemically evoked activity (T1.3); (3) the number of spikes in the second 3 sec period of evoked activity (T4.6). Since it is possible that the resting level of activity could be viewed as a variable baseline, or noise, against which the superimposed sensory signals are compared by the organism, it was thought that a measure of the difference between the resting and evoked activity might provide a single index containing information of possible biological utility. Thus the difference between the prior resting level (B) and the total amount of activity evoked by chemical stimulation in each 3 sec period was calculated for each presentation by subtracting B from TI_ 3 and T4_6, respectively. These difference measures are termed D1. 3 and D4. 6 . In order to compare and display amounts of response in different experiments and load conditions, the response measures were normalized by assigning the response to the last preload presentation of each stimulus a value of 10. These normalized measures are termed B', T'I_3, T'4_6, D'l_3, and D'4. 6. Percent changes in response magnitude from the preload state to the loaded state were calculated from the medians of the preload and load response magnitudes. The probability values of the changes in response magnitude from preload to load were calculated by means of the Wilcoxon Matched-Pairs Signed-Rank Test. The probability values for differences in magnitude changes within the load conditions were calculated by the Mann-Whitney U Test [26]. RESULTS Data derived from 25 samples of second-order neural activity gathered before and after gastric loading in 15 rats are reported in this paper. There were 15 samples of preload and load measurement of chemically evoked activity under conditions of simple distention (air load). The other 10 samples were taken from trials in which 0.3 M NaC1 was intubed into the stomach; in one of these trials no oral stimuli were presented as an indication of oral stimulation-independent changes in resting activity.
Effect o f Gastric Loads on O.1 M NaCl-Evoked Activity The effects of gastric loads of both air and 0.3 M NaC1 on the neural responses to 0.1 M NaC1 are graphically represented in Fig. 1 for the D'4_ 6 time period. Each point
563
on this composite graph represents the median normalized activity measure calculated from all trials in which 0.1 M NaC1 was presented during that time period. An estimate of variability in each time period is provided by the inclusion of the interquartile range (IQR). The slowly developing response decrement after gastric loading seen in Fig. 1 is typical of the decremental pattern seen in the other measurements (i.e., T ' l _ 3 , T ' 4 _ 6 , D ' l _ 3 ) . The simultaneous consideration of resting (B measures) and total evoked activity (T measures) provided by the D measures emphasized the decrement seen in the T measures alone. This emphasis was not due to a consistent increase in the resting activity after gastric loading. The possible contributions of a decremental trend over time during the preload period were examined for their possible effects on the results. Although slight negative slopes were seen with both T' 1_3 (a slope of - 0 . 0 5 ) and T ' 4 _ 6 (a slope of - 0 . 0 2 ) , only in the former case could the null hypothesis be rejected that the slope was not significantly different from zero (p
Gastric Modulation of Gustatory Afferent Activity' JOHN F. GLENN 2 AND ROBERT P. ERICKSON a
Duke University (Received 5 June 1975)
GLENN, J.F. AND R.P. ERICKSON. Gastric modulation of gustatory afferent activity. PHYSIOL. BEHAV. 16(5) 561-568, 1976. - The neural activity evoked by stimulation of the rat's tongue with NaCl, sucrose, HCI, and QHC1 was recorded from 25 small populations of second-order gustatory neurons in the nucleus of the solitary tract, both before and after gastric loading. A slowly developing (several minutes) load-induced depression in the response evoked by the taste solutions was noted, with no significant differences seen between loading with air or 0.3 M NaCl. Differences were evident in the magnitude of the decrement for the various stimuli; the order of decrement was sucrose > NaCl > HC1;the response to QHCI showed no clear decrement. The effect of gastric loading, as tested with air loads, is reversible. The magnitude of the load effect was seen to be less with greater levels of food deprivation. The results of this study therefore, demonstrate the existence of gastric effects on the reactivity of second-order gustatory afferents to chemical stimulation of the rat's tongue. Taste Gastric distention Multineuronal activity
Nucleus of the solitary tract
Food intake
Afferent interaction
factors, determine the organism's feeding response to a foodstuff [321. Given that sensory information from the oral cavity influences the acceptance and rejection of foodstuffs, the question remains - what are the mechanisms by which a given oral stimulus may result in different behaviors? It is possible that, under the influence of both humoral and neural factors, the central integrative, perhaps hypothalamic, mechanisms may simply send a stop or go signal, or graded amounts of stop and go signals, to the mot or centers in the medulla subserving ingestive behavior. However, it is also possible that the hypothalamus and other CNS structures may exert some effect on the afferent input instead of, or in addition to, a more direct effect on the efferent m o t o r system. While such central influences might act on sensory receptors or relays to bias or modify at some level the oropharyngeal sensory input to the feeding reflex, it is also possible that these central influences may be augmented or supplanted by more direct influences exerted on the oral sensory input by ascending visceral afferents from the gut. The juxtaposition of the terminals of afferents for vagally mediated interoceptive sensory information and sensory information from the oropharyngeal cavity in the nucleus tractus solitarius of the hindbrain strongly suggests such direct influences. There are several experiments which demonstrate that
ALTHOUGH feeding behavior historically has been studied from several perspectives, the emphasis of modern research originated within the framework of homeostatic regulatory mechanisms. In particular, the thoughts of Claude Bernard on the constancy of the internal environment have pointed the directions for research on the bases of homeostatic regulation. The rhythmic fluctuations seen in meal frequency, meal size, and the foodstuffs selected consequently are seen as behavioral compensations for deviations from certain criteria of normality in the milieu interieur. While internal, postingestional (PI) signals of b o t h gastric and humoral origin are generally accepted as providing interoceptive information about the intraorganic state, relatively little systematic consideration has been given to the role of external, oropharyngeal factors, especially taste and olfaction, within the mechanisms involved in the regulation of food intake. Yet, it is obvious that the mechanisms controlling ingestive behavior are as inextricably bound to external sensory factors as to internal factors. As examples, the recovery of feeding behavior in brain-damaged animals has been shown to be tastedependent (hypothalamic hyperphagics and aphagics; see [28] ), and satiety has been shown to be sensory specific [5, 14, 15]. Thus, there are both internal (PI) and external (oral and olfactory) classes of situational input which, in conjunction with innate predispositions and experiential
1This paper is based on a doctoral dissertation submitted at Duke University by the first author. This work was supported in part by NSF Grant GB 33464 and in part by NIMH Predoctoral Fellowship 1S01-MH-58159-01. 2j. F. Glenn is presently serving as a Captain in the U.S. Army Medical Service Corps and is assigned to the Behavioral Research Directorate of the U.S. Army Human Engineering Laboratory at Aberdeen Proving Ground, MD 21005. 3Send reprint requests to R. P. Erickson, Professor of Psychology and Associate Professor of Physiology at Duke University, Durham, NC 27706. 561
562
GLENN AND ERICKSON
postingestionally initiated control of the afferent message exists in the gustatory system. Behavioral evidence of postingestional changes in gustatory reactivity has been gathered in both humans [33 ] and dogs [5 ]. Electrophysiological evidence for changes in chemically evoked activity after stomach distention in toads and frogs has been demonstrated in the first-order gustatory afferents of the glossopharyngeal nerve [2,10] and in the gustatory receptor potential [24]. While the mechanisms underlying these behavioral and electrophysiological effects are not clear, it seems obvious that the gustatory message may be altered in some manner by changes in the milieu interieur. Although there is behavioral evidence from humans and dogs for modulation of gustatory afferent information, there has been no electrophysiological verification of intragastrically induced neural changes in the taste system of mammals. Since a large portion of the literature on feeding behavior involves work with rats, it was thought that a demonstration of sensory modulation in that animal would be useful. The nucleus solitarius of the medulla, with its multiple special and general visceral afferent input, would seem to provide a logical locus for integration of visceral and oral information. It is the purpose of this study, therefore, to investigate the effect of intragastric loading on the chemically evoked activity of second-order neurons of the nucleus tractus solitarius of the rat. METHOD
Animals Fifteen female Sprague-Dawley albino rats (Charles River), ranging in weight from 185 to 245 g, were used. Thirteen of the animals were isolated before surgery without access to food, but with ad lib water, for varying periods of time. The two animals not isolated were taken directly from the colony cage, where they had been maintained on an ad lib schedule of both food and water intake. Surgery. The animals were prepared for surgery under a combination of anesthetic (inhalation and barbituate) and neuroleptanalgesic (narcotic analgesic and major tranquilizer) techniques. The animals were first lightly sedated with Fluothane (halothane, Ayerst). Upon sedation, atropine sulfate was given (0.3 mg/kg IP). A single injection of Brevital Sodium (sodium methohexital-Lilly), a rapidly effective, ultra-shortacting barbituate, was used to induce anesthesia (25 mg/kg IP). This dosage provided a surgical level of anesthesia for about 15 min. The trachea was cannulated with polyethlene tubing, and a size 8 Bardic infant intra-gastric feeding tube (C. R. Bard Inc.) was inserted into the esophagus until the tip of the tube passed into the stomach. The animal was then mounted into a head holder which allowed free access to the oral cavity [8]. Innovar Injection (McNeil: narcotic analgesic-fentanyl; neuroleptic-droperidol) was administered (0.1 mg/kg IM) and Anucaine (Calvin Chemical), a long-lasting local anesthetic, was injected in small amounts into the cut edges of the wound to provide a surgical level of anesthesia. The surgical techniques for the exposure of the medulla were carried out under a Zeiss operating microscope by first removing portions of the occipital skull, and then aspirating the cerebellum to expose the floor of the fourth ventricle. The EKG and body temperature were monitored throughout the procedure, and body temperature was maintained between 35 ° and 37.5°C by means of an
electric body warmer. The animal was immobilized with Flaxedil (gallamine triethiodide, Davis and Geck; 60 mg/kg IP) and was artificially respirated by means of a Palmer or Harvard respirator pump. Pneumothorax was performed when necessary to reduce brain movement.
Procedure Recording. The chemically evoked responses in small populations of second-order neurons of the Nucleus Tractus Solitarius were recorded with tungsten microelectrodes (Transidyne General 404-20). The identification of secondorder gustatory activity was made according to the methods and criteria of Doetsch and Erickson [6]. The active electrode was positioned with a three-coordinate micromanipulator, the vertical movements of which were hydraulically driven. A bare nichrome wire positioned on the floor of the fourth ventricle with a second micromanipulator acted as the second recording electrode. Leads from the two electrodes, as well as the animal ground lead, were connected to a Grass P-15 preamplifier. The preamplifier output was fed to a Grass RM-122 low-level preamplifier, the output of which was connected in parallel to an audio monitor, a Tektronix 502 dual-beam oscilloscope, and a magnetic tape recorder (Sony 650-4 or Teac 2340). Stomach loading. The intragastric feeding tube, which was inserted into the cardiac portion of the stomach, was connected to a 10 ml syringe. In order to allow introduction of both chemical and inert loads in the same preparation, simple distention of the stomach was accomplished by the introduction of a volume of air, rather than the more widely used procedure of an intragastric balloon. The peculiarity of the anatomy of the rat's cardiac sphincter, which prevents emesis, probably contributed to the tightness of seal between the intragastric tube and sphincter, and allowed retention of the inert air load throughout the course of the load condition. Surgical exposure of the stomach was made in several rats in a preliminary investigation, and visual inspection showed that this procedure of air loading was reliable and repeatable in that air did not leak out, but could be withdrawn simply and easily by pulling back on the plunger of the syringe. In the volumes injected, the area of distention in these animals seemed to be well confined to the stomach, with no noticeable duodenal distention. The volume injected in the air load condition was approximately 8 ml (range 6 - 1 0 ml). The solution chosen for chemical loading was 0.3 M NaC1. This concentration was deemed appropriate in that it was sufficiently concentrated to remain largely in the stomach for several minutes with reduced clearance relative to water [ 12] without being pathological. The amounts of NaC1 intubed were slightly less than those of air (range 5 . 5 - 8 . 5 ml). Visual inspection of the medulla showed no noticeable movement of the medulla relative to the recording electrode during distention, nor did the neural activity at this time suggest movement. Data collection. Chemical solutions of 0.1 M NaC1, 0.03 M HC1, 0.01 M QHC1, and 1.0M Sucrose were prepared in deionized water, and were introduced by means of a gravity flow system into a chamber fitted around the anterior portion of the tongue. A stimulus presentation consisted of a flow of solution over the tongue for approximately 1 0 - 1 5 sec. Each presentation was followed
GASTRIC MODULATION OF GUSTATORY A F F E R E N T ACTIVITY by a 4 5 - 6 0 sec continuous rinse with deionized water, except in Experiment 1-1 where rinse time was approximately 20 sec. In 14 cases, presentations were made before, during and after removal of the stomach load as a check for the reversibility of distention effects. Data analysis. The records of neural activity consisted of populations of unit responses in which one or several single units were resolvable. The data were thus amenable to analysis by means of a spike counter with a variable threshold amplitude discriminator circuit; the output of the amplitude discriminator was in the form of square wave pulses. For further analysis, the output of the spike counter in each 500 msec period was printed on paper tape by a Franklin Electronics counter. Three measures of activity for each presentation of a stimulus were calculated from the printed spike counts: (1) the average rate of resting (spontaneous) activity during the 3 sec preceding each presentation (B); (2) the number of spikes in the first 3 sec of chemically evoked activity (T1.3); (3) the number of spikes in the second 3 sec period of evoked activity (T4.6). Since it is possible that the resting level of activity could be viewed as a variable baseline, or noise, against which the superimposed sensory signals are compared by the organism, it was thought that a measure of the difference between the resting and evoked activity might provide a single index containing information of possible biological utility. Thus the difference between the prior resting level (B) and the total amount of activity evoked by chemical stimulation in each 3 sec period was calculated for each presentation by subtracting B from TI_ 3 and T4_6, respectively. These difference measures are termed D1. 3 and D4. 6 . In order to compare and display amounts of response in different experiments and load conditions, the response measures were normalized by assigning the response to the last preload presentation of each stimulus a value of 10. These normalized measures are termed B', T'I_3, T'4_6, D'l_3, and D'4. 6. Percent changes in response magnitude from the preload state to the loaded state were calculated from the medians of the preload and load response magnitudes. The probability values of the changes in response magnitude from preload to load were calculated by means of the Wilcoxon Matched-Pairs Signed-Rank Test. The probability values for differences in magnitude changes within the load conditions were calculated by the Mann-Whitney U Test [26]. RESULTS Data derived from 25 samples of second-order neural activity gathered before and after gastric loading in 15 rats are reported in this paper. There were 15 samples of preload and load measurement of chemically evoked activity under conditions of simple distention (air load). The other 10 samples were taken from trials in which 0.3 M NaC1 was intubed into the stomach; in one of these trials no oral stimuli were presented as an indication of oral stimulation-independent changes in resting activity.
Effect o f Gastric Loads on O.1 M NaCl-Evoked Activity The effects of gastric loads of both air and 0.3 M NaC1 on the neural responses to 0.1 M NaC1 are graphically represented in Fig. 1 for the D'4_ 6 time period. Each point
563
on this composite graph represents the median normalized activity measure calculated from all trials in which 0.1 M NaC1 was presented during that time period. An estimate of variability in each time period is provided by the inclusion of the interquartile range (IQR). The slowly developing response decrement after gastric loading seen in Fig. 1 is typical of the decremental pattern seen in the other measurements (i.e., T ' l _ 3 , T ' 4 _ 6 , D ' l _ 3 ) . The simultaneous consideration of resting (B measures) and total evoked activity (T measures) provided by the D measures emphasized the decrement seen in the T measures alone. This emphasis was not due to a consistent increase in the resting activity after gastric loading. The possible contributions of a decremental trend over time during the preload period were examined for their possible effects on the results. Although slight negative slopes were seen with both T' 1_3 (a slope of - 0 . 0 5 ) and T ' 4 _ 6 (a slope of - 0 . 0 2 ) , only in the former case could the null hypothesis be rejected that the slope was not significantly different from zero (p
