Anthony Sclafani: Consummate scientist.

In this article we review the scientific contributions of Anthony Sclafani, with specific emphasis on his early work on the neural substrate of the ventromedial hypothalamic (VMH) hyperphagia-obesity syndrome, and on the development of diet-induced obesity (DIO). Over a period of 20 years Sclafani systematically investigated the neuroanatomical basis of the VMH hyperphagia-obesity syndrome, and ultimately identified a longitudinal oxytocin-containing neural tract contributing to its expression. This tract has since been implicated in mediating the effects of at least two gastrointestinal satiety factors. Sclafani was one of the first investigators to demonstrate DIO in rats as a result of exposure to multiple palatable food items (the "supermarket diet"), and concluded that diet palatability was the primary factor responsible for DIO. Sclafani went on to investigate the potency of specific carbohydrate and fat stimuli for inducing hyperphagia, and in so doing discovered that post-ingestive nutrient effects contribute to the elevated intake of palatable food items. To further investigate this effect, he devised an intragastric infusion system which allowed the introduction of nutrients into the gut paired with the oral intake of flavored solutions, an apparatus her termed the "electronic esophagus". Sclafani coined the term "appetition" to describe the effect of intestinal nutrient sensing on post-ingestive appetite stimulation. Sclafani's productivity in the research areas he chose to investigate has been nothing short of extraordinary, and his studies are characterized by inventive hypothesizing and meticulous experimental design. His results and conclusions, to our knowledge, have never been contradicted.

Randall Sakai, chronic social stress, and the research tradition of Curt Richter.

This paper describes Randall Sakai's professional career from graduate school at the University of Pennsylvania, through postdoctoral work at Rockefeller University, and to being an independent investigator at the University of Cincinnati. He was fortunate in having Alan Epstein, Bruce McEwen, and Eliot Stellar as mentors. Early in Sakai's graduate work, Epstein and Stellar introduced him to Curt Richter, the legendary investigator at Johns Hopkins. This early introduction to Richter and his tradition of research was crucial for Sakai's scientific development. We review Sakai's research with the Visible Burrowing System (VBS) at Cincinnati. This was the most original of Sakai's research interests. His experimental proficiency in the investigation of salt appetite, food intake, and obesity led him to focus on the effect of chronic social stress on food intake, body composition, metabolism, and the distribution of fat. He and his colleagues, many of them his students, were able to demonstrate that chronic social stress produced changes in metabolism and fat distribution that were characteristic of an incipient metabolic syndrome that could lead to obesity. This did not solve the problem, but showed the way to further investigation. This opening up of problems to experimental investigation was a hallmark of Richter's research. Thus, Sakai worked in the mainstream of the research tradition of Richter. He did what he revered.

Adolf Meyer's Influence on Curt Richter: Selection, Support, and Mentoring.

Adolph Meyer influenced Curt Richter from the time Richter became a graduate student in Psychology at Johns Hopkins in 1919 until Meyer retired in 1941. Meyer was on Richter's thesis committee. After Richter received his degree, Meyer selected him to replace J.B. Watson. During the next 20 years, Meyer gave Richter strong support in terms of equipment, laboratory space for animal research, and opportunities to teach medical students, attend clinical rounds, and do clinical research. It is less well known that Meyer also mentored Richter's scientific and professional development. Meyer's mentoring was so successful that Richter adopted the major scientific ideas of Meyer, especially psychobiology, distrust of theory, and respect for experiment. Thus, Meyer's ideas became the framework for Richter's famous research program that produced his major discoveries of behavior exerting homeostatic controls, psychoendocrinology, and biological clocks.

Hervey, Harris, and the parabiotic search for lipostatic signals.

This paper is an introduction to the papers by Hervey and Harris that describe their experimental use of parabiosis in rats and mice to search for circulating lipostatic signals. Beginning in 1959 with Hervey's foundational paper, they detected three parabiotic signals: the Hervey signal decreases food intake and fat mass in rats; the antilipogenic factor (ALF) decreased fat mass, but not food intake in rats; and the leptin-dependent signal in lean partners of ob/ob mice decreased fat mass, but not food intake. The known lipostatic signals, leptin and insulin, have been candidates for the Hervey and ALF signals, but insulin has been excluded and the evidence for leptin is inconclusive. The site of production of the three parabiotic signals and their molecular structure are not known and specific mechanisms of their lipostatic control are incompletely understood. Given their potential importance for understanding the physiology of lipostatic controls and for developing new therapies for obesity, Hervey and Harris make a strong argument for further research on the three parabiotic signals.

Synergistic relationship between the Columbia University Appetitive Behavior Seminar and the satiating effect of cholecystokinin.

Synergism between the Columbia University Appetitive Behavior Seminar and the research program of Smith and Gibbs on the satiating effect of cholecystokinin during the past 40 years is described. The Seminar was synergistic with the research program in five ways. First, the steady parade of speakers gave us a window on the varied and interesting work going on in the field. Second, the Seminar was the kind of audience for presentations of the work-in-progress on CCK that scientists hope for and rarely find. Criticism by members of the Seminar was relentless and constructive, and ideas for further experiments or new ways to tackle problematic data poured forth. Third, members of the Seminar did experiments that facilitated the experimental success of the research program. Fourth, members of the Seminar tutored us on topics that we wanted to import into the research program on CCK. Fifth, and probably most important, members of the Seminar gave us the encouragement, good humor, and friendship so necessary for coping with the struggles of the scientific life.

Hippocampal neurons inhibit meal onset.

There is extensive research regarding the neural mechanisms involved in satiety and meal termination; in contrast, there is very limited understanding of how meal onset is regulated. On the basis of several converging lines of evidence, we hypothesized that hippocampal neurons form a memory of a meal and inhibit meal onset during the postprandial period. As a first step, we tested whether reversible inactivation of the hippocampus with muscimol infusions after the end of one meal would accelerate the onset of the next meal. To test this, adult male Sprague-Dawley rats (N = 23) were implanted with a cannula aimed at the right or left dorsal hippocampus and then trained to consume a 32% sucrose solution at a scheduled time daily. On the experimental day, hippocampal neuronal activity was temporarily disrupted during the postprandial period by infusing muscimol (0.5 µg/µl; 1 µl) 5 min after the rats stopped consuming the sucrose solution. Compared to vehicle infusions, muscimol infusions significantly decreased the latency to start the postinfusion meal and increased the size of the postinfusion meal. In addition, muscimol disrupted the relationship between the size of a meal and length of the following postprandial period. These effects of muscimol on meal onset were not due to an effect on the speed of consumption. Collectively, these findings are consistent with the hypothesis that hippocampal neurons suppress meal initiation during the postprandial period. Given that overeating can impair hippocampal function, these findings suggest that impaired hippocampal functioning is a cause and consequence of overeating and obesity.

Stephen C. Woods: a precocious scientist.

To investigate the early scientific development of Steve Woods, I reviewed his research during the first decade after he received his doctoral degree in 1970. The main parts of his research program were conditioned insulin secretion and hypoglycemia, Pavlovian conditioning of insulin secretion before a scheduled access to food, and basal insulin as a negative-feedback signal from fat mass to the brain. These topics were pursued with experimental ingenuity; the resulting publications were interesting, clear, and rhetorically effective. Although the theoretical framework for his experiments with insulin was homeostatic, by the end of the decade he suggested that classic negative-feedback homeostasis needed to be revised to include learning acquired by lifestyle. Thus, Woods functioned as a mature scientist from the beginning of his research-he was very precocious. This precocity also characterized his teaching and mentoring as recalled by two of his students during that time, Joseph Vasselli and Paul Kulkosky. The most unusual and exemplary aspect of his precocity is that the outstanding performance of his first decade was maintained during the subsequent 30years.

The inhibitory potency of SCH 23390 and raclopride on licking for sucrose increases across brief-access tests.

There is extensive pharmacological and microdialysis evidence that central dopamine mechanisms are important for the mediation of the rewarding and reinforcing functions of sweet taste. One aspect of the pharmacological evidence for dopaminergic mediation of sweet reward is unclear. That is the positive interactive effect of ingestive experience and DA antagonist treatment reported by Wise and his colleagues in 1978 [1]. They showed that the inhibitory potency of pimozide increased over repetitive tests of saccharin (0.1%) ingestion. When pimozide was given before 8 daily, 10-minute tests in rats licking [2] or lever pressing [3] for 32% sucrose, however, the inhibitory effect of pimozide did not increase across tests. To reinvestigate the problem, we used a computer-assisted, repetitive, brief-access technique [4, 5] in which 10 male, non-deprived, Sprague Dawley rats licked 0.5M sucrose for 60s in four trials with a 30-second intertrial interval. Thirty minutes before the first trial, each rat received an ip injection of the D1 antagonist SCH 23390, the D2/3 antagonist raclopride, or vehicle. SCH 23390 and raclopride decreased licking significantly, their inhibitory effects increased significantly within and across the 4 trials, and the temporal pattern of their inhibitory effects on latencies, and on cumulative and total licks was different. Thus we confirm an increase of the inhibitory potency of DA antagonists across ingestive tests and show for the first time that the interaction differs between D1 and D2/3 antagonists.

The conditioned satiating effect of orosensory stimuli.

Prior to the introduction of sham feeding as a method for studying the controls of meal size, the dominant view was that gustatory stimulation activated the ingestion of palatable diets and postingestional stimulation inhibited it. Early sham feeding studies with rats challenged this view because they showed that, contrary to expectation, rats did not eat continuously the first time they were given a sham feeding test. They ate a larger meal than when tested under normal conditions but stopped eating and showed all the signs of satiety soon after. Only after two or more sham feeding tests did they eat continuously. Subsequent research, reviewed here, established that experience ingesting a diet under real feeding conditions leads to the development of a classically conditioned form of satiation based on an association between gustatory stimulation and some consequence of gastrointestinal stimulation by the ingested food. This conditioned orosensory satiating effect extinguishes when sham feeding occurs repeatedly without intervening real feeding tests. Thus gustatory stimulation both stimulates and inhibits meal size. An experimental implication of this finding is that intake during sham feeding must be shown to be maximal before sham feeding can be used to measure only the orosensory stimulation of the diet. Another implication is that the analysis of a change in meal size produced by some treatment should now include measurement of the potency of the conditioned orosensory satiating effect as well as the potencies of orosensory stimulation and postingestive negative feedback.

Unacknowledged contributions of Pavlov and Barcroft to Cannon's theory of homeostasis.

Cannon's theory of homeostasis is the first, major, American contribution to physiological thought. Although it is clear that Cannon's account of homeostasis is personal and based primarily on the work of his laboratory, Cannon made it easy for readers to mistake his 1929 paper and 1932 book for a comprehensive review of the literature relevant to homeostasis. This is unfortunate because Cannon never acknowledged the important contributions of two of his contemporaries, Ivan Pavlov and Joseph Barcroft. Since he did not mention them, their contributions are rarely discussed. This paper attempts to correct this historical problem in two ways. First, I describe the unacknowledged contributions of Pavlov and Barcroft. Then I consider the possible reasons why Cannon ignored them.

Introduction to four papers on Curt Richter and analysis of his scientific practice.

Curt Richter was one of the founders of our field. He did outstanding research for over 60 years. Richter did fundamental work in appetite for food and minerals in the 1930s and discovered the homeostatic functions of ingestive behavior. This paper introduces four papers on specific topics of his work by contemporaneous experts. Each of the papers reviews Richter's experiments and then shows how the problem developed since he left it. The papers demonstrate that providing the historical basis for contemporary science is not only instructive, it is also heuristic for the science waiting to be done. In addition to introducing the four papers, I analyze the scientific ideas, values, and men that influenced Richter's scientific practice. I conclude that Claude Bernard, Walter Cannon, Francois Magendie, and Maurice Arthus were important for Richter's scientific ideas and values, but it was the joy of research that explains his experimental success for over 60 years.

Gastric preloads of corn oil and mineral oil produce different patterns of increases of c-Fos-like immunoreacitve cells in the brain of 9-12 day-old rats.

Equivolumetric gastric preloads of corn oil and mineral oil administered to rats on postnatal day 12 (P12) inhibited intake equally during a 30-min test of independent ingestion (II), but preloads of corn oil inhibited intake significantly more than preloads of mineral oil on P15 and P18 [Weller, A., Gispan, I.H., Armony-Sivan, R., Ritter, R.C., Smith, G.P., 1997. Preloads of corn oil inhibit independent ingestion on postnatal day 15 in rats. Physiol. Behav. 62, 871-874]. It is possible that the equivalent inhibition of intake by the oil preloads on P12 resulted from the failure of the preabsorptive sensory properties of the preloads to be discriminated by peripheral or central sensory mechanisms. To investigate this possibility, we administered equivolumetric gastric preloads of 25% corn oil and 25% mineral oil to pups on P9-12 and counted the number of c-Fos-like immunoreactive (CFLI) cells in central sites that are activated by food intake and postingestive preabsortive mechanisms in adult rats and in pups on P10-11. The major result was that preloads of 25% corn oil and 25% mineral oil that produced equivalent inhibition of II intake produced differential increases of CFLI cells in the forebrain and hindbrain. Specifically, preloads of corn oil increased the number of CFLI cells in the caudal Nucleus Tractus Solitarius significantly more than preloads of mineral oil. Furthermore, preloads of corn oil increased the number of CFLI cells in the Paraventricular and Supraoptic nuclei, but preloads of mineral oil did not. This differential pattern of increases of CFLI cells is evidence that the brain discriminates the preabsorptive sensory properties of preloads of corn oil and mineral oil on P9-12.

Ontogeny of ingestive behavior.

Review of the ontogeny of the controls of independent ingestion reveals that some of the direct and indirect controls of meal size identified in adult rats function in the first three postnatal weeks. The controls appear sequentially and some of them change their potency after they emerge. Indirect controls exerted by metabolism and adiposity do not emerge until the fourth postnatal week or later in the postweaning period. Recent experiments in rats with monogenic obesities involving the leptin and cholecystokinin receptors have demonstrated the usefulness of independent ingestion in the detection of the earliest expression of hyperphagia. Although much remains to be learned about the normal controls of independent ingestion, it is clear that it provides relevant information about the development of normal and abnormal controls of meal size in rodents that is useful for translational research into the controls of meal size in normal and obese children.

Cholecystokinin-8 increases Fos-like immunoreactivity in myenteric neurons of the duodenum and jejunum more after intraperitoneal than after intravenous injection.

The objective of this study was to measure the relative efficacy and potency of cholecystokinin-8 (CCK-8) given by intraperitoneal (i.p.) and intravenous (i.v.) injection to stimulate Fos-like immunoreactivity (Fos-LI) in neurons of the myenteric plexus in the duodenum and jejunum. The subjects for his experiment were 40 male Sprague-Dawley rats divided into eight treatment groups (n=5 rats per treatment). Four groups of rats were injected with 5, 10, and 40 microg/kg sulfated CCK-8 and saline (control) i.p., and the remaining groups with the same treatments i.v. We then detected Fos-LI, a marker for neuronal activation, in the myenteric plexus of the duodenum and jejunum, in response to the previous doses and routes. All of the CCK-8 doses administered by both routes increased Fos-LI in the myenteric plexus of the duodenum and jejunum significantly more than saline did. Although both routes were efficacious in increasing Fos-LI, CCK-8 i.p. was significantly more potent than CCK-8 i.v. These data provide immunohistochemical evidence that i.p. administration of CCK-8 is a more potent stimulant of Fos-LI in the neurons of the myenteric plexus of the duodenum and jejunum than i.v. injection.

Accumbens dopamine mediates the rewarding effect of orosensory stimulation by sucrose.

To investigate the hypothesis that central dopamine mediates the rewarding effect of orosensory stimulation by sucrose that increases eating, specific dopamine antagonists were administered prior to sham-feeding tests in which rats ingested various concentrations of sucrose. Sham feeding (SF) was used to preserve the orosensory stimulation of sucrose while it eliminated sucrose's postingestive effects. The antagonists decreased intake as a function of the concentration of sucrose. Microstructural analysis of licking demonstrated that the antagonists did not affect the motor performance of licking. Thus, the inhibitory effect of the antagonists on intake was due to a decrease in the rewarding effect of sucrose. Microdialysis experiments demonstrated that dopamine was released in the nucleus accumbens during SF and that the synaptic action of dopamine in the accumbens was necessary for the normal rewarding effect of orosensory stimulation by sucrose.

Oral sucrose stimulation increases accumbens dopamine in the rat.

Although taste can influence meal size and body weight, the neural substrate for these effects remains obscure. Dopamine, particularly in the nucleus accumbens, has been implicated in both natural and nonnatural rewards. To isolate the orosensory effects of taste from possible postingestive consequences, we investigated the quantitative relationship between sham feeding of sucrose and extracellular dopamine in the nucleus accumbens with microdialysis in rats. Sucrose intake linearly increased as a function of concentration (0.03 M, 18.07 +/- 2.41 ml; 0.1 M, 30.92 +/- 2.60 ml; 0.3 M, 43.28 +/- 2.88 ml). Sham feeding also stimulated accumbens dopamine overflow as a function of sucrose solution concentration (0.03 M, 120.76 +/- 2.6%; 0.1 M, 140.28 +/- 7.8%; 0.3 M, 146.27 +/- 5.05%). A second experiment used the same protocol but clamped the amount of sucrose ingested and revealed a similar, concentration-dependent dopamine activation in the nucleus accumbens. This is the first demonstration of a quantitative relationship between the concentration-dependent rewarding effect of orosensory stimulation by sucrose during eating and the overflow of dopamine in the nucleus accumbens. This finding provides new and strong support for accumbens dopamine in the rewarding effect of sucrose.

Decreased hypothalamic concentration of neuropeptide Y correlates with onset of hyperphagia in fa/fa rats on postnatal day 12.

An increased action of hypothalamic neuropeptide Y (NPY) has been proposed as a major factor in the pathophysiology of the obesity syndrome in Zucker (fa/fa) rats. Using a developmental strategy to test this hypothesis, we showed previously that significantly more arcuate NPY was expressed in fa/fa pups than in lean littermates on postnatal day (P) 2 and throughout the preweaning period [Physiol. Behav. 67 (1999) 521], and that hyperphagia first appeared on P12 [Am. J. Physiol. 275 (1998) R1106]. To test the hypothesis further, we used a specific radioimmunoassay to measure the concentration of hypothalamic NPY peptide in lean (+/+ and +/fa) and obese fa/fa Zucker rat pups on P9, P10, and P12. The concentration of NPY in fa/fa pups was not significantly different from that of the other genotypes. There was, however, a significant decrease in NPY concentration from P9 to P12 in fa/fa pups, but not in lean pups. The combination of increased NPY message and decreasing concentration of NPY peptide in fa/fa pups with age is consistent with, but does not prove, increased release of hypothalamic NPY in fa/fa pups just before and on P12 when hyperphagia emerges. These results provide further support for the importance of hypothalamic NPY in the phenotypic expression of hyperphagia in the fa/fa pups during the second postnatal week.