Margery C. Beinfeld

The distribution of cholecystokinin immunoreactivity in the central nervous system of the rat as determined by radioimmunoassay.

The regional distribution of cholecystokinin (CCK) in the rat brain was determined utilizing a radioimmunoassay which detects both gastrin and CCK. CCK concentration is highest in the caudate nucleus (10-14 ng CCK 8 equivalents/mg protein), followed by the cerebral cortex. Within the cerebral cortex, CCK is highest in the cingulate, pyriform, and entorhinal areas. There are substantial CCK concentrations in all other brain regions except pons, medulla and cerebellum. CCK is widely distributed in the hypothalamus, where it is highest in the median eminence and ventromedial nucleus. Considerable CCK-like immunoreactivity is also present in the posterior lobe of the pituitary gland, but is not detectable in anterior and intermediate lobes. Though the antisera used in this study cross-react with gastrin the dominant CCK-like material found in rat brain co-elutes with sulfated CCK 8 and separates from gastrin on Sephadex G-25 and HPLC chromatography.

Cholecystokinin innervation of the ventral striatum: A morphological and radioimmunological study

Immunocytochemistry, radioimmunological assay after surgical cuts, anterograde degeneration and retrograde tracing of fluorescent dyes were used in order to elucidate the cholecystokinin-containing afferents to the ventral striatum (nucleus accumbens, olfactory tubercle and ventral part of the caudate-putamen). In agreement with the report by Hökfelt et al., midbrain cholecystokinin-containing cells supply the posteromedial parts of the nucleus accumbens and olfactory tubercle, as well as the subcommissural part of caudate-putamen. Brainstem cholecystokinin afferents also reach more rostral parts of the ventral striatum including the rostrolateral olfactory tubercle. The ascending cholecystokinin axons enter the medial forebrain bundle at the meso-diencephalic border and maintain a rough medial to lateral topography at the caudal diencephalon. A second major cholecystokinin pathway, with possible origin in the piriform and medial prefrontal cortices and/or the amygdala, projects to the subcommissural caudate-putamen, the olfactory tubercle, the lateral part of the nucleus accumbens and the dorsal part of the bed nucleus of stria terminalis. Finally, the rostral part of the dorsal caudate-putamen receives a substantial cholecystokinin innervation from the basolateral amygdala and possibly from the neocortex. According to radioimmunological data, the descending telencephalic cholecystokinin system accounts for about 60% of all cholecystokinin in the rostral forebrain. The combined use of morphological and biochemical methods provided evidence for a partially overlapping distribution and possible interaction between an ascending brainstem and descending telencephalic cholecystokinin fiber systems within the striatum and related rostral forebrain areas.

Cholecystokinin in the central nervous system: a minireview

This review focuses on the structure, distribution, neuronal pathways, receptor binding, release, biosynthesis and degradation of CCK in the central nervous system. Other aspects of the isolation and chemistry of CCK (1), its role in satiety (2), as a hormone or neurotransmitter (3,4), and its evolution (5) have been reviewed recently.

Demonstration of a putative receptor site for cholecystokinin in rat brain

Abstract High affinity (apparent K d ⋍ 0.65 nM) and saturable (B max ⋍ 6.6 fmoles/mg protein) binding sites for cholecystokinin (CCK) have been demonstrated in crude synaptosomal membranes from rat cerebral cortex. Scatchard analysis of the equilibrium binding data indicates a single population of non-interacting sites. The specific binding of 125 I-CCK 33 to brain membranes is reversible ( t 1 2 ⋍ 6 minutes ), and can be inhibited by structurally related CCK analogues but not by unrelated neuropeptides or drugs. Specific CCK binding reaches equilibrium at a temperature-dependent rate, and is abolished when membranes are pre-treated with heat. Kinetic and competition studies suggest that these high affinity binding sites represent physiologically-relevant receptors for CCK in brain.

Evidence for defective mesolimbic dopamine exocytosis in obesity-prone rats

The association between dietary obesity and mesolimbic systems that regulate hedonic aspects of feeding is currently unresolved. In the present study, we examined differences in baseline and stimulated central dopamine levels in obesity‐prone (OP) and obesity‐resistant (OR) rats. OP rats were hyperphagic and showed a 20% weight gain over OR rats at wk 15 of age, when fed a standard chow diet. This phenotype was associated with a 50% reduction in basal extracellular dopamine, as measured by a microdialysis probe in the nucleus accumbens, a projection site of the mesolimbic dopamine system that has been implicated in food reward. Similar defects were also observed in younger animals (4 wk old). In electrophysiology studies, electrically evoked dopamine release in slice preparations was significantly attenuated in OP rats, not only in the nucleus accumbens but also in additional terminal sites of dopamine neurons such as the accumbens shell, dorsal striatum, and medial prefrontal cortex, suggesting that there may be a widespread dysfunction in mechanisms regulating dopamine release in this obesity model. Moreover, dopamine impairment in OP rats was apparent at birth and associated with changes in expression of several factors regulating dopamine synthesis and release: vesicular monoamine transporter‐2, tyrosine hydroxylase, dopamine transporter, and dopamine receptor‐2 short‐form. Taken together, these results suggest that an attenuated central dopamine system would reduce the hedonic response associated with feeding and induce compensatory hyperphagia, leading to obesity.—Geiger, B. M., Behr, G. G., Frank, L. E., Caldera‐Siu, A. D., Beinfeld, M. C., Kokkotou, E. G., Pothos, E. N. Evidence for defective mesolimbic dopamine exocytosis in obesity‐prone rats. FASEB J. 22, 2740–2746 (2008)

Selective depletion of somatostatin in rat brain by cysteamine

Abstract A single injection of cysteamine (300 mg/kg, subcutaneously) results in a 70–80% decrease in somatostatin levels in the periventricular nucleus where somatostatin-producing neurons are located and the median eminence where somatostatinergic nerve terminals are. The drug seems quite selective: no changes in levels of other neuropeptides — LH-RH, vasopressin, enkephalin, VIP, CCK — were observed in the same animals.

Distribution of cholecystokinin (CCK) in the hypothalamus and limbic system of the rat

Abstract Cholecystokinin (CCK) concentrations were determined in microdissected individual nuclei of the hypothalamus and limbic system using a specific and sensitive CCK radioimmunoassay (RIA). The CCK levels in the hypothalamus were highest (> 2 mg CCK8 sulfate equivalents/mg protein) in the ventromedial, dorsomedial, periventricular, arcuate nuclei, the median eminence, and mamillary body. Most regions of the limbic system had higher levels of CCK than the hypothalamus and in particular the CA3 region of the hippocampus, the lateral septal nucleus and the medial and lateral amygdaloid nuclei had levels of CCK which were similar to some cerebral cortical areas. This study confirms previous preliminary CCK distribution studies in the rat and indicates that CCK in addition to its wide distribution in the cerebral cortex, is present in abundance in many areas of the hypothalamus and limbic system.

CCK-containing terminals in the hippocampus are derived from intrinsic neurons: an immunohistochemical and radioimmunological study

Cholecystokinin (CCK) was originally detected and sequenced as a 33 amino acid peptide (CCK3a) in gastrointestinal tissuea,5, 8. Subsequently, a CCK-like substance was detected in the brains of several vertebrate species2, 3,6,tz which was found to share the same 8 amino acid C-terminal sequence (CCKs) as CCKa3 a. In the rat brain, large amounts of CCK are present in nerves in both cortical and subcortical regions 6,1°,12. Particularly high concentrations are found in the hippocampus, as demonstrated by radioimmunoassay 1. lmmunohistochemical techniques have demonstrated the presence of both CCK-containing somata and terminals in the hippocampus 4,7. Because other brain regions such as the septum and entorhinal cortex, which project to the hippocampus, also contain high concentrations of CCK 1, the CCK-containing terminals in the hippocampus may be derived from either the intrinsic CCK neurons or from extrinsic CCK neurons, or from a combination of both. The present investigation therefore had two goals. First, an analysis of the distribution of CCK-containing cell bodies and terminals in the hippocampus was undertaken, using immunohistochemical techniques. Second, the source of the CCKcontaining terminals was determined, by isolating the hippocampus from its afferent projection areas by radio frequency lesions and then measuring the CCK content of the hippocampus by radioimmunoassay. As previously reported, the rabbit CCKantisera used in these experiments cross-react with gastrin. The dominant CCK-like material found in rat brain, however, coelutes with. sulfated CCKs and separates from gastrin on G-25 sephadex and high pressure liquid chromatography 1. Twenty-six male albino rats, weighing 250-350 g, were used for these experiments. Three were used for immunohistochemistry, 24 for the hippocampal deafferen-

Prejunctional and postjunctional effects of neuropeptide Y at the noradrenergic neuroeffector junction of the perfused mesenteric arterial bed of the rat

Summary: The effect of neuropeptide Y (NPY) on periarterial nerve stimulation-induced release of norepinephrine (NE) and increase in perfusion pressure in the perfused mesenteric arterial bed of the rat was examined. Perfusate effluents were continuously collected and assayed for endogenous NE by high-pressure liquid chromatography (HPLC) coupled to electrochemical detection. Perfusion pressure was continuously monitored by means of a pressure transducer. Periarterial nerve stimulation (8 or 16 Hz, 60 V, 2-ms duration for 30 s) resulted in a readily detectable increase in NE release and perfusion pressure that was attenuated by the prior administration of tetrodotoxin (TTX) (10−5 M) or guanethidine (5 × 10−5 M). NPY exerted both prejunctional and post-junctional effects on noradrenergic neurotransmission in this preparation. The peptide produced a concentration-dependent reduction in the release of NE over a concentration range of 10−10-10−7 M. A similar inhibition effect occurred at 8. 10. and 16 Hz. In contrast, low concentrations (10−10 and 10−9 M) decreased the effect of nerve stimulation on perfusion pressure, whereas higher concentrations (10−7 M) produced a marked potentiation. The α2-adrenoceptor antagonist, yohimbine, did not alter the inhibitory effect of NPY on evoked NE release or the effect on perfusion pressure. Prazosin similarly did not alter the inhibitory effect of NPY on NE release but prevented the increase in perfusion pressure. We conclude that NPY modulates noradrenergic neurotransmission in the mesenteric arterial bed by decreasing the evoked release of NE and producing a concentration-dependent bi-phasic response on vascular smooth muscle. The inhibition of NE produced by low concentrations of NPY results in a decrease in perfusion pressure whereas high concentrations produce a marked increase in perfusion pressure.

Distribution of cholecystokinin (CCK) in the rat lower brain stem nuclei

The cholecystokinin (CCK) concentration in individual brain stem nuclei of rat was determined using the Palkovits punch method19 and the CCK RIA3, CCK has a unique distribution in the brain stem, unlike other neuropeptides and biogenic amines8,19. In general, the CCK levels in the brain stem are 5-20% of rat cerebral cortex. The colliculi, midbrain central gray, nucleus of the solitary tract, and the interpeduncular nucleus had the highest CCK content (2.7-1 ng CCK mg protein).