Does the angle of trocar insertion affect the fascial defect caused? A porcine model.

INTRODUCTION: With an incidence of 0-5.2%, trocar site hernias frequently occur following laparoscopy. It is unclear to what extent the angle of trocar insertion affects the size of the fascial defect caused. Hence, we performed a porcine model. METHODS: In October 2022, a total of five female pigs were euthanized. In alternating order, three bladeless and two bladed conical 12-mm trocars were inserted at an angle of 45° on each side for 60 min twice each pig. For this purpose, an epoxy resin handmade cuboid with a central channel that runs at an angle of 45° was used. Subsequently, photo imaging and defect size measurement took place. The results were compared with those of our previously conducted and published porcine model, in which the trocars were inserted at an angle of 90°. Effects of trocar type (bladed vs. bladeless) and angle on defect size were analyzed using a mixed model regression analysis. RESULTS: The bladeless trocars caused statistically significant smaller defects at the fascia than the bladed (23.4 (SD = 16.9) mm2 vs. 41.3 (SD = 14.8) mm2, p < 0.001). The bladeless VersaOne trocar caused the smallest defect of 16.0 (SD = 6.1) mm2. The bladed VersaOne trocar caused the largest defect of 47.7 (SD = 10.5) mm2. The defect size of the trocars used at a 45° angle averaged 30.5 (SD = 18.3) mm2. The defect size of trocars used at a 90° angle was significantly larger, averaging 58.3 (SD = 20.2) mm2 (p = 0.007). CONCLUSION: When conical 12-mm trocars are inserted at a 45° angle, especially bladeless ones, they appear to cause small fascial defects compared with insertion at a 90° angle. This might lead also to a lower rate of trocar hernias. Bladeless trocars might cause smaller fascial defects than bladed trocars.

How often is prophylactic parastomal mesh placement performed after rectal resection without sphincter preservation? An analysis of German nationwide hospital discharge data among 41,697 patients.

PURPOSE: The European Hernia Society guidelines of parastomal hernias, published in 2017, strongly recommend prophylactic synthetic non-absorbable mesh upon the construction of a permanent end colostomy to reduce the incidence of parastomal hernias. This study aims to evaluate the implementation of the guidelines in Germany. METHODS: This is a retrospective multicentric analysis conducted in December 2022 at the University Hospital Brandenburg an der Havel. Anonymous data on rectal resection without sphincter preservation in the period 2010-2020 were extracted from the German nationwide hospital discharge data set. Individuals with a hernia and < 18 years old were excluded. Another exclusion criterion was a performed colectomy or proctocolectomy with an ileoanal pouch and placement of an absorbable mesh. The primary endpoint was the annual rate of prophylactic parastomal mesh placement following rectal resection without sphincter preservation in Germany. Cases reporting both non-absorbable mesh placement and rectal resection without sphincter preservation were considered prophylactic mesh insertions. RESULTS: A total of 41,697 patients received a rectal resection without sphincter preservation and without non-absorbable mesh placement. Among these individuals, 27,089 were male and 14,608 were female. The rate of reoperations (3.1%) and the length of hospital stay (25.3 days ± 19.32) remained almost constant during these 10 years. The rate of prophylactic mesh placement was increasing from 0.2% (n = 8) in 2010 to 6.4% (n = 198) in 2020. CONCLUSIONS: Currently, only the minority of patients who have undergone rectal resection without sphincter preservation receive prophylactic mesh insertion.

4-Acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid inhibits adrenocorticotropin secretion from anterior pituitary cells.

Mouse clonal ACTH-secreting corticotrophs (AtT-20 cells) possess a membrane Ca2+-activated Cl- conductance which is partially blocked by the disulfonic stilbene derivative 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS). In the current study the effect of SITS on the ACTH secretory process was evaluated. SITS markedly blocked basal and forskolin-stimulated ACTH secretion from AtT-20 cells (IC50 = 2.7 x 10(-4) M). Both CRF-induced ACTH secretion and forskolin-stimulated GH secretion from acutely dispersed rat anterior pituitary cells were inhibited by SITS (IC50 = 2.4 and 1.3 x 10(-4) M, respectively). SITS did not alter unstimulated or forskolin-elicited cAMP synthesis in AtT-20 cells, and in fact, could inhibit ACTH secretion in response to cAMP-independent agonists such as the calcium channel activator BAY-K-8644 or the protein kinase-C activator 12-tetradecanoyl-phorbol-13-acetate (IC50 = 2.6 and 2.4 x 10(-4) M, respectively). SITS did not alter the secretion of amylase from isolated exocrine pancreatic acinar cells. Its action was also fully reversible; after its removal from the incubation medium, cells secreted ACTH without a change in response to forskolin activation. Increasing extracellular Ca2+ or the addition of up to 10(-3) M tetraethylammonium or 4-aminopyridine did not reverse the inhibitory pattern of SITS action, suggesting that its inhibitory effect is most likely not due to hyperpolarization of AtT-20 cell membranes. The inability of amiloride to inhibit ACTH secretion further suggests that inhibition of ACTH secretion provoked by SITS is not due to a blockade of Cl-/HCO3- exchange. On the other hand, SITS was able to block 44% of basal 36Cl uptake by AtT-20 cells. Exchange of incubation medium chloride for gluconate or a reduction in the osmotic strength of the medium reduced both basal and secretagogue-stimulated ACTH secretion. The data suggest that SITS may modulate chloride-dependent, osmotically driven secretion from AtT-20 cells.

Internalization and subcellular distribution of radiolabeled somatostatin-28 in mouse anterior pituitary tumor cells.

The ACTH-secreting mouse AtT-20/D16-16 (AtT-20) tumor corticotroph possesses receptors for the tetradecapeptide somatostatin (S-14) to which the 14-amino acid N-terminal extension somatostatin-28 (S-28) also binds. When AtT-20 cells are exposed to either S-14 or S-28 for extended periods of time, a marked decrease in S-14 receptor density is observed. Since receptor down-regulation is frequently associated with internalization of ligand and/or receptor, the present study was designed to establish whether AtT-20 cells could in fact internalize S-28 and to determine the subcellular localization of internalized peptide. Cells were incubated in the presence of [Leu8, D-Trp22,125I-Tyr25] S-28 for 1, 4, and 18 h; washed with PBS; and harvested. Cell pellets were fixed, sectioned, and analyzed by light and electron microscopic autoradiography. Uptake of radiolabeled S-28 (90% of all cells) was inhibited by 80% when unlabeled S-14 was coincubated with [125I]S-28. Of the cell compartments examined, plasma membrane, secretory granules, lysosomes, Golgi apparatus, and nuclear membrane all had distinct time-dependent labeling patterns. Plasma membranes were maximally labeled 1 h after exposure to [125 I] S-28. Secretory granule and lysosomal labeling was observed within 1 h, but was maximal after 18 h of exposure. Labeling of the granule compartment preceded that of the Golgi apparatus. The nuclear compartment (membranes plus nuclei) was also labeled significantly after 18 h of incubation. However, the nuclear membrane itself was labeled after only 1 h of exposure to the ligand. The data suggest that radiolabel is transferred from the plasma membrane to the intracellular organelles as a function of exposure time. Labeling of the secretory compartment suggests that granules may bind and protect internalized peptide from lysosomal degradation. Appearance of label in the nuclear compartment suggests that S-28 (and S-14) may have effects on transcriptional activity in AtT-20 cells.

Somatostatin-14 and somatostatin-28 pretreatment down-regulate somatostatin-14 receptors and have biphasic effects on forskolin-stimulated cyclic adenosine, 3',5'-monophosphate synthesis and adrenocorticotropin secretion in mouse anterior pituitary tumor cells.

Activation of somatostatin-14 (S-14) receptors on mouse AtT-20 pituitary tumor cells by S-14 or somatostatin-28 (S-28) inhibits forskolin-stimulated cAMP synthesis and ACTH secretion. In this study, the effects of prolonged exposure of cells to S-14 or S-28 was found to reduce, in a time- and concentration-dependent fashion, the density of S-14 receptors without affecting the affinity of these sites for [125I]Tyr11-S-14. This response was rapidly reversible after removal of peptide from incubation media. Additionally, S-14 and S-28 pretreatment also resulted in a time-dependent sensitizing effect on forskolin-stimulated cAMP formation and ACTH secretion which preceded S-14 receptor down-regulation. Enhancement of the forskolin response was concentration dependent, with maximal effects observed at 10(-8) M with either peptide. Higher pretreatment concentrations of S-14 resulted in an abolition of the enhanced biological response to forskolin; pretreatment with S-28 (10(-6) M) depressed forskolin- and (-)isoproterenol-induced cAMP formation below levels observed in nonpretreated cells. The enhancing effect of S-14 and S-28 required new protein synthesis, since it was partially blocked by cycloheximide; the depressor effect was independent of new protein synthesis. Both the enhanced and depressed forskolin responses after peptide pretreatment were reversible after withdrawal of S-14 or S-28; normalization of the forskolin response (cAMP formation and ACTH secretion) followed the return to control levels of S-14 receptor density. Pretreatment of cells with 10(-8) M or 10(-6) M S-28 increased and decreased, respectively, the ACTH secretory response to agonists which act in the absence of prior cAMP synthesis such as 8-bromo-cAMP, A-23187, and phorbol ester. The data suggest that S-14 receptor down-regulation is not causally associated with the sensitizing effects of S-14 and S-28 on adenylate cyclase and that the S-14 receptor may be also coupled to other effector systems which are involved in regulating the secretory function of AtT-20 cells.

Relationship between receptor binding and biopotency of somatostatin-14 and somatostatin-28 in mouse pituitary tumor cells.

Somatostatin-14 (S-14) acts via specific receptors to inhibit basal as well as hormone- and forskolin-stimulated ACTH secretion in tumor cells (AtT-20/D16-16) of mouse anterior pituitary. In addition S-14 inhibits the stimulated but not basal cAMP accumulation. The potency of somatostatin-28 (S-28) for regulating these processes in these tumor cells has not been reported. In this study we have investigated the relationship between receptor-binding affinities of S-14 and S-28 and their biopotency in these cells. Membrane receptors for S-14 characterized using [125I-Tyr11]S-14 as the radioligand [maximum binding capacity (Bmax) = 1.28 +/- 0.1 pmol/mg; dissociation constant (Kd) = 1.1 +/- 0.04 nM] bound S-28 with 3-fold greater affinity than S-14. Binding sites quantitated using an S-28 analog [Leu8, D-Trp22, 125I-Tyr25]S-28 as radioligand (Bmax = 1.18 +/- 0.15 pmol/mg; Kd = 0.08 +/- 0.06 nM) also exhibited greater affinity for S-28 than S-14. Forskolin-stimulated cAMP accumulation and ACTH secretion in these cells were inhibited to a greater extent (4- and 9-fold, respectively) by S-28 than S-14. Preincubation of the cells with S-14 and S-28 (10(-7) M) resulted in a marked decrease (36% and 71%, respectively) of S-14 receptor concentration. Coincubation of the cells with both S-14 and S-28 led to 56% decrease in S-14 receptor binding. The responsiveness of the cells to forskolin stimulation of ACTH secretion and cAMP accumulation was significantly enhanced by preincubation with S-14 (10(-7) M) whereas the responsiveness to forskolin was completely abolished by preincubation with S-28. Simultaneous exposure of the cells to both S-14 and S-28 resulted in a partial reversal of the inhibiting effect of S-28 on forskolin-stimulated cAMP accumulation in these cells but did not result in a partial reversal of the inhibitory effect of S-28 on forskolin-stimulated ACTH secretion in these cells. These results demonstrate that S-28 is more potent than S-14 in AtT-20/D16-16 cells, its greater potency arising from its greater affinity for binding to S-14 receptors. The differential effects of these peptides after preincubation on the responsiveness of ACTH secretion and cAMP accumulation in these cells to forskolin stimulation suggests the possibility of existence of distinct S-14 and S-28 receptors, but these could not be identified by direct binding experiments using the S-14 and S-28 analogs employed in the study as radioligands.(ABSTRACT TRUNCATED AT 400 WORDS)

Is atrial natriuretic peptide synthesized and internalized by gonadotrophs?

The present study was designed to determine whether atrial natriuretic peptide (ANP) could be both synthesized and internalized by rat anterior pituitary gonadotrophs. ANP synthesis was assessed by in situ hybridization of ultrathin frozen sections of anterior pituitary to a biotinylated 30-base oligonucleotide to rat ANP mRNA. As revealed by the immunogold technique, only gonadotrophs were labeled by the probe. At the subcellular level, ANP mRNA was observed at both the nuclear and cytoplasmic levels in gonadotrophs, and labeling of the latter compartment was quantitatively more intense. Internalization of ANP was investigated by an in vivo ultrastructural autoradiographic approach. Intravenous injection of [125I]ANP resulted in rapid labeling within 1 min of the plasma membrane, cytoplasmic matrix, secretory vesicle, and mitochondrial compartments and the Golgi apparatus; these compartments were labeled throughout the remainder of the time course studied (1-30 min). Peak labeling of the plasma membrane compartment was at 1 min and diminished from that point; labeling in the Golgi apparatus peaked 5 min postinjection, while in the other compartments labeling was fairly uniform over the time course. The lysosomal compartment was also radiolabeled; however, only 2 and 5 min after injection of [125I]ANP. The findings demonstrate that gonadotrophs can both synthesize and internalize extracellular ANP. These observations can be extended to suggest that ANP has both autocrine and paracrine actions in the anterior pituitary gland. Since the peptide neither stimulates nor antagonizes the release of any anterior pituitary hormone, these actions are probably unrelated to the adenohypophyseal secretory function.

Y-1 adrenocortical tumor cells contain atrial natriuretic peptide receptors which regulate cyclic nucleotide metabolism and steroidogenesis.

The effects of atrial natriuretic peptide (ANP) on adrenocortical fasciculata cells were examined in the ACTH-responsive Y-1 mouse adrenocortical tumor cell line. Y-1 cell membranes rapidly bound [125I]ANP, with equilibrium binding (22 C) reached within 45 min. Binding of [125I]ANP was inhibited in a concentration-dependent manner by unlabeled ANP and atriopeptin-I (IC50, approximately 1.2 X 10(-9) and 1.6 X 10(-8) M, respectively), but not by C- or N-terminal-deleted ANP fragments, ACTH, or arginine vasopressin (up to 10(-6) M). Scatchard analysis revealed a single class of high affinity binding sites with a Kd of 1.6 X 10(-10) M and a binding capacity of 560 fmol/mg protein. Photo-affinity labeling demonstrated the specific binding of ANP to two protein entities of 130 and 63 kDa. ANP stimulated both cGMP synthesis and secretion from Y-1 cells (EC50, approximately 3.5 X 10(-9) M). Release of the nucleotide was inhibited by probenecid (IC50, approximately 5 X 10(-5) M). The atrial peptide partially inhibited ACTH-stimulated cAMP formation (IC50, approximately 10(-8) M) and partially antagonized basal and ACTH-stimulated steroidogenesis. The data demonstrate the presence in Y-1 cells of specific and saturable ANP receptors, activation of which leads to changes in cyclic nucleotide metabolism and inhibition of steroidogenesis.

Dopamine inhibits prolactin secretion stimulated by the calcium channel agonist Bay-K-8644 through a pertussis toxin-sensitive G protein in anterior pituitary cells.

In primary culture of anterior pituitary cells, BAY-K-8644, a calcium channel agonist, stimulated PRL secretion by 83% with EC50 of 18 nM. This effect was blocked by nifedipine, a calcium channel antagonist. The stimulations of PRL secretion induced by potassium (50 mM) and BAY-K-8644 were additive. Dopamine inhibited basal as well as BAY-K-8644-stimulated PRL secretion by 64% and 75%, respectively, and with respective EC50 values of 4.5 and 0.6 nM. In the presence of 50 mM K+, dopamine only partially blocks the dose-dependent stimulation of PRL secretion induced by the calcium channel agonist. The inhibitory dopamine effect was blocked by (+)butaclamol, a specific dopamine receptor antagonist. The dopamine response was also blocked by 1-sulpiride, a specific dopamine D2 receptor antagonist, and mimicked by RU 24926, a specific dopamine D2 receptor agonist, suggesting that the dopamine effect on BAY-K-8644-stimulated PRL secretion was mediated through a D2 dopamine receptor. Although unknown, the mechanism by which dopamine inhibited the BAY-K-8644-stimulated PRL secretion involves a GTP binding protein sensitive to Bordetella pertussis toxin. In fact, the dopamine inhibition of PRL secretion induced by the calcium channel agonist was blocked by the pretreatment of cells with the toxin. These results suggest that dopamine D2 receptors in lactotroph cells modulate calcium influx through a GTP binding protein.

Synthesis, internalization, and localization of atrial natriuretic peptide in rat adrenal medulla.

Some, though not all studies, have indicated that atrial natriuretic peptide (ANP) can bind to adrenal medullary cells. ANP-like immunoreactivity (ANP-LI) has also been identified in catecholamine-secreting cells. Together, these findings suggest that ANP may be taken up and/or synthesized in the adrenal medulla. The present study was designed to ascertain, by in situ hybridization, whether adrenal chromaffin cells could synthesize ANP, to define by an in vivo ultrastructural autoradiographic approach, whether ANP could, in fact, bind to rat adrenal medulla cells, to determine whether there was a cellular [noradrenaline (NA) vs. adrenaline (A)] selectivity in the binding process, and to establish whether extracellular [125I]ANP could be internalized by these cells. The cellular and subcellular distribution of endogenous ANP-LI was also investigated in both cell types by cryoultramicrotomy and immunocytochemical approaches. The in situ hybridization studies indicate the presence of mRNA to ANP in about 15% of adrenal medullary cells. Intravenous injection of [125I]ANP resulted in a 3-fold, preferential and specific radiolabeling of A-as compared to NA-containing cells. In A-containing cells, plasma membranes were significantly labeled 2 and 5 min post injection; cytoplasmic matrix, mitochondria, and secretory granules throughout the time course studied (1-30 min post injection). Lysosomes, rough endoplasmic reticulum, Golgi apparatus, and nuclei were not labeled. ANP-LI was identified in both NA- and A-containing cells; in the former, it was almost exclusively localized in secretory vesicles, in the latter it was detected in plasma membranes, cytoplasmic matrix, nuclear euchromatin, some mitochondria and relatively fewer granules than in NA-containing cells. The findings suggest that ANP may be synthesized primarily in NA-containing cells and that A-containing cells primarily bind and internalize the extracellular (endogenous or exogenous) atrial peptide. The data suggest that ANP secreted by adrenal medullary chromaffin cells may have distal paracrine actions or interactions with coreleased catecholamines and neuropeptides. Binding and internalization may reflect an action of ANP on the secretory function of A-containing cells.

Calcium efflux and secretion of alpha-amylase from rat pancreas.

1 The efflux of (45)Ca from rat pancreas was investigated in relation to the secretion of alpha-amylase.2 The (45)Ca space in the pancreas was increased 1.6 times when pancreatic tissue was incubated in media in which the extracellular calcium concentration was reduced from 1.0 to 0.05 mM.3 Carbachol increased the rate of (45)Ca efflux from the tissue and this effect was associated with an increase in the release of alpha-amylase. Dibutyryl cyclic adenosine monophosphate (AMP) also evoked a secretory response, but did not alter the rate of (45)Ca efflux. In combination with carbachol, the dibutyryl analogue of cyclic AMP reduced the carbachol-stimulated increase in (45)Ca efflux while enhancing the release of secretory protein.4 The stimulatory effect of carbachol on (45)Ca efflux was observed when pancreatic tissues were incubated in media containing a high concentration of ethyleneglycolbis (beta-aminoethyl)-N,N'-tetraacetic acid (EGTA).5 Atropine blocked the effects of carbachol on both (45)Ca efflux and secretion of alpha-amylase.6 It was concluded that intracellular calcium can and may sustain stimulus-secretion coupling in the rat exocrine pancreas.

The possible role of histamine in the coritsone induced gastric acid hypersecretion of the guinea-pig.

1. The effects of cortisone treatment combined with agents influencing histamine metabolism were studied on the free and total acid output in pylorus ligated guinea-pigs.2. It was found that reduction in the synthesis of histamine produced by the administration of alpha-methylDOPA significantly inhibited the acid response to cortisone.3. Enhancement of the oxidative deamination of histamine brought about by the administration of diamine oxidase or heparin inhibited cortisone induced acid hypersecretion significantly.4. Inhibition of the oxidative deamination of histamine by aminoguanidine and iproniazid resulted in a significant increase of the cortisone induced acid hypersecretory response.5. Inhibition of the methylation of histamine by chlorpromazine or 1,4-methylhistamine inhibited the cortisone induced hypersecretory response significantly.6. Studies with labelled histamine indicated that cortisone increases the sequestration of histamine to the stomach.

Corticotropin releasing factor stimulation of protein carboxylmethylation in mouse pituitary tumor cells.

A putative role for the protein carboxylmethylase (PCM) enzyme has been suggested in exocytotic secretion. The involvement of 3H-methyl incorporation into protein carboxylmethyl esters during corticotropin releasing factor (CRF)-induced ACTH secretion from AtT-20/D16-16 mouse pituitary cells was investigated. Protein carboxylmethylation and ACTH secretion both increased as a function of extracellular CRF concentration, and both processes were temporally parallel up to 60 min incubation. The less potent [Met(O)21]-CRF also stimulated increases in protein carboxylmethylation and ACTH secretion. The free acid analogue of CRF did not alter either process. A combination of the PCM inhibitors, 3-deazaadenosine and L-homocysteine thiolactone, reduced both CRF-stimulated protein carboxylmethylation and ACTH release. Dexamethasone, known to inhibit ACTH secretion and synthesis, inhibited both CRF-stimulated protein carboxylmethylation and ACTH secretion.

PHI stimulates ACTH release from pituitary tumor cell.

The identification of PHI (the 27-amino acid peptide (P) having an N-terminal histidine (H) and a C-terminal isoleucine amide (I] in median eminence suggested that PHI influences the secretory function of the anterior pituitary. The effects of PHI on ACTH release from clonal mouse pituitary corticotrophs were investigated. The secretory response to PHI was correlated with a prior increase in cyclic AMP accumulation. Both cyclic AMP synthesis and ACTH secretion were increased by PHI in a concentration-dependent manner. PHI was a less effective agonist of cyclic nucleotide synthesis and ACTH secretion than VIP. The secretory response to PHI was blocked by the calcium channel antagonist, nifedipine, and by dexamethasone. Somatostatin and oxotremorine blocked both PHI-stimulated cyclic AMP formation and ACTH secretion. The observation that VIP in high concentrations can elicit ACTH secretion from normal rat anterior pituitary suggested that a VIP-like substance may modulate ACTH secretion. However, the finding that PHI does not elicit ACTH release from primary cultures of dispersed anterior pituitary, coupled to its relatively lower potency compared to VIP, indicate that corticotrophs are not an important target for PHI.

Atrial natriuretic factor does not affect basal, forskolin- and CRF-stimulated adenylate cyclase activity, cAMP formation or ACTH secretion, but does stimulate cGMP synthesis in anterior pituitary.

The report that ANF inhibits basal and CRF-stimulated adenylate cyclase activity in anterior pituitary homogenates suggested that the atrial peptide could inhibit ACTH secretion. This possibility was investigated in the ACTH-secreting AtT-20 mouse pituitary tumor cell line as well as homogenates or primary cell cultures from rat anterior hypophysis. ANF (up to 5 X 10(-7) M) was found to be completely ineffective in stimulating basal, CRF- and/or forskolin-stimulated adenylate cyclase activity, cAMP accumulation and ACTH secretion. Similarly, ANF had no effect on spontaneous or GRF-induced GH release from cells in primary culture. ANF receptors, however, are present in AtT-20 cells and anterior pituitary cells as evidenced by the ability of the peptide to stimulate intracellular cGMP accumulation. The data, therefore, suggests that ANF does not have a negative modulatory action on the secretory function of anterior pituitary. The role of cGMP in any other action(s) of ANF remains unknown.

Synthesis and internalization of atrial natriuretic factor in anterior pituitary cells.

Atrial natriuretic factors (ANF) are a family of peptides originally identified in atrial cardiocytes and having natriuretic, diuretic and vasorelaxatory properties. ANF was recently reported to be synthesized in anterior pituitary cells. In the current study, the cell-specific sites of binding and internalization of ANF in the adenohypophysis, a site of ANF action, were investigated. By in vitro autoradiographic techniques, [125I]ANF was found to bind specifically to both anterior and posterior lobes of the pituitary, but not to the intermediate lobe. 5 min following intravenous injection of [125I]ANF, radiolabel was detected by ultrastructural autoradiography on gonadotrophs, corticotrophs and lactotrophs, but not thyrotrophs or somatotrophs. Label was found at both the plasma membrane level and cytoplasmic matrix, but not in the nuclei, of all three cell types. The cellular and subcellular localization of endogenous ANF-like immunoreactivity was also investigated. Specific immunoreactivity was also detected in gonadotrophs, corticotrophs and some lactotrophs. At the subcellular level in all three cell types, immunoreactivity was localized in the cytoplasmic matrix, on secretory granule membranes, and in mitochondria. Less dense staining was observed in nuclear euchromatin; endoplasmic reticulum and Golgi apparatus were not labelled in any of the cells. The data suggest that notwithstanding their reported ability to synthesize ANF, exogenous ANF can also bind and be internalized by adenohypophyseal gonadotrophs; the binding and presence of endogenous ANF-like immunoreactivity in corticotrophs and lactotrophs suggest that these cells may be sites of action for the atrial peptide.

Evidence that AVP receptors in AtT-20 corticotrophs are not coupled to secretion of POMC-derived peptides.

This study reports the presence in AtT-20 corticotrophs of high affinity-low capacity receptors for arginine-vasopressin (AVP), whose binding capacity was considerably enhanced by the divalent metal ion nickel. These binding sites, when analyzed in the presence of nickel, showed high affinity for AVP, vasotocin and oxytocin, but recognized to a lesser extent the V2-agonist 1-deamino-AVP, as well as V1-antagonists. Surprisingly, AVP failed to alter secretion of proopiomelanocortin (POMC)-derived peptides from the cells or corticotropin-releasing factor (CRF)-induced cAMP synthesis, as reported in normal corticotrophs. Exposure of cells to CRF elicited an increase in mRNAPOMC levels, while, in contrast, AVP was without significant effect. It thus appears that in AtT-20 tumor cells, the AVP receptors are not coupled to either the biochemical or biological cellular response.

Immunocytochemical localization, binding, and effects of atrial natriuretic peptide in rat adipocytes.

The metabolic effects of atrial natriuretic peptide (ANP) have not been widely investigated. Since adipocyte cells represent a model system extensively used to examine the metabolic actions of many peptide hormones, we sought to establish whether ANP could bind to adipocyte membranes, alter cyclic nucleotide metabolism, and affect spontaneous or hormone-stimulated lipolysis. Using in vitro autoradiographic techniques, radiolabelled ANP was found to bind specifically to mammary gland fat cells. Additionally, endogenous ANP-like immunoreactivity could be localized in the plasma membrane compartment and cytoplasmic matrix of fat cells, but not in fat vacuoles. [125I]ANP bound to single high affinity sites (Kd = 0.72 nM) in fat cell membranes. The binding was rapid (equilibrium within 1 min at 25 degrees C) and specific. The atrial peptide was capable of stimulating a time- and concentration-dependent increase in cGMP accumulation in isolated adipocytes, but had no effect on spontaneous or stimulated [-)-isoproterenol, ACTH, forskolin) cAMP formation. ANP did not alter the increase in glycerol production stimulated by l-epinephrine in isolated fat cells. While i.v. infusion of ANP stimulated a marked increase in circulating levels of cGMP, the atrial peptide did not alter plasma triglyceride levels. These data demonstrate the presence of specific ANP binding sites on adipocyte membranes and internalization of ANP-associated immunoreactivity. These receptors are biochemically functional given the ability of ANP to augment cGMP formation. The peptide, however, does not exert an action on adipocyte lipolysis. Adipocytes, therefore, represent an ANP target tissue in which the physiological action of the peptide is yet to be defined.

Psychologic assessment of morbidly obese patients undergoing gastric bypass: a comparison of preoperative and postoperative adjustment.

Thirty-three morbidly obese patients underwent gastric bypass operation after intensive medical and psychiatric evaluation. Five psychometric tests were administered before and after operation. The study patients were found to have less self-esteem and more depressive traits than a normal population. This did not change after operation despite weight loss. High levels of optimism were not associated with better weight loss. However, patients who understood before operation that the success of the operation depended upon changing their eating behavior lost more weight. After operation patients expressed satisfaction with life and a new freedom from constant hunger. There was a reported decrease in organic symptoms, an increase in social activities, an improvement in interpersonal relationships, and a social usefulness not experienced previously. In selected patients with no active psychiatric disease or psychologic instability, gastric bypass, coupled with consistent postoperative reinforcement, produces behavioral changes that can lead to permanent weight loss without concomitant psychologic deterioration.

Atrial natriuretic factor-induced cGMP accumulation in rat anterior pituitary cells in culture is not coupled to hormonal secretion.

While atrial natriuretic factor (ANF) does not influence ACTH secretion, it was reported to have a marked stimulatory effect on the intracellular accumulation of cGMP in rat anterior pituitary cells in culture. Since many biological actions of ANF appear coupled to its excitatory action on target cell guanylate cyclase, the current study was designed to characterize the ANF-induced cGMP response in anterior pituitary with a view to determining whether the nucleotide plays a regulatory role in the secretory function of this gland. A 3 min exposure of cells in primary culture to 300 nM ANF (99-126) or 100 microM sodium nitroprusside (SNP), a stimulator of guanylate cyclase, causes maximal 10- and 3-fold elevations of cGMP levels, respectively. Following a progressive decrease, 6- and 2-fold increases over basal cGMP levels are still observed after 180 min of incubation with ANF (99-126) and SNP, respectively. The half-maximal stimulation of cGMP accumulation induced by a 10 min exposure to ANF (99-126), or rat atriopeptin II (ANF 103-125) is observed at 9 +/- 2 and 125 +/- 22 nM, respectively. ANF fragments (99-109) and (111-126), as well as human cardiodilatin (hANF 1-16), do not alter cGMP levels. Basal and ANF-induced cGMP levels are at least 10-fold higher in cell populations enriched in gonadotrophs compared to gonadotroph-impoverished preparations. A 3 h incubation of cells with ANF (0.1-1000 nM), however, fails to modify spontaneous or LHRH-induced LH secretion. Similarly, ANF does not alter spontaneous release of GH, TSH or PRL. The data suggest indirectly that gonadotrophs represent a principal site at which ANF acts to stimulate cGMP synthesis, but that the nucleotide is not a specific regulator of the LH secretory process; nor is it generally involved as a second messenger in the secretory function of any cell type of the anterior pituitary gland.