exocrine pancreatic function during cold blockade of

Experimental Physiology (1990), 75, 401-406 Printed in Great Britain

EXOCRINE PANCREATIC FUNCTION DURING COLD BLOCKADE OF THE VAGUS IN CHRONIC EXPERIMENTS ON CALVES ROMUALD ZABIELSKI, PAWEL PODGURNIAK, STEFAN GRZEGORZ PIERZYNOWSKI AND WIESLAW BAREJ Department of Animal Physiology, Faculty of Veterinary Medicine, Warsaw Agricultural University, SGGW-AR Nowoursynowska 166, 02-766 Warsaw, Poland

(MANUSCRIPT RECEIVED I NOVEMBER 1988, ACCEPTED 6 DECEMBER 1989)

SUMMARY

Four calves were prepared surgically to investigate the exocrine pancreatic function in chronic experiments. Cooling devices were implanted on both vagi for temporary, reversible, thermal blockade of the conductivity in the nerves. Cooling of the vagi caused significant decreases of volume, total protein content and trypsin activity in pancreatic juice. All of these variables recovered to the control level immediately after the cooling. Results presented indicate an important role of vagally driven information on the exocrine pancreatic secretion in calves. INTRODUCTION

The role of CNS and vago-vagal (long) reflexes in controlling pancreatic exocrine secretion have been intensively studied, but much remains unclear. The effect of section of the vagus nerves on the response of the pancreas to secretin has produced conflicting results, from an unchanged response or a reduced response to no response at all (for references see Grundy, Hutson & Scratcherd, 1983). The contribution of the vagus nerves to the control of pancreatic secretion varies from species to species (for references see Harada, Nakagawa & Kato, 1982 and Singer, 1987). In an attempt to differentiate between vagal and hormonal control, both pharmacological and nerve section have been used. The effects of truncal vagotomy have been reported by many authors (Thomas, 1950 for early references; Tankel & Hollander, 1958; Magee, Fragola & White, 1965; Henriksen, 1969; Moreland & Johnson, 1971; Konturek, Becker & Thompson, 1974; Debas, Konturek & Grossman, 1975; Grundy et al. 1983). In addition, pancreatic secretion has been studied after complete extrinsic denervation brought about by autotransplantation (Wang & Grossman, 1951; Singer, Solomon & Grossman, 1980; Kohler, Nustede, Barthel & Schafmayer, 1987). Acute vagotomy is a once and for all act and only one observation is possible in one animal. Cold block allows repeated interruptions to the vagal supply of the pancreas with complete recovery on every occasion. Nevertheless, any type of acute interruption in the vagal supply could produce a sudden imbalance between cholinergic, adrenergic and peptidergic control mechanisms. Chronic vagotomy on the other hand allows the establishment of adaptive processes which can occur even in the absence of any apparent structural changes within the intrinsic nerve plexuses of the pancreas (Radke & Stach, 1986). This present paper extends the information available concerning the role of the vagus nerves with respect to the interdigestive exocrine pancreatic function in another species, the ruminant calf. Downloaded from Exp Physiol (ep.physoc.org) by guest on July 10, 2011

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Animals and surgery Four Friesian male calves,1-5-2 months old, 45-60 kg body weight (BW) were used. The calves fasted for 24 h prior to surgery and operations were done using xylazine neuroleptoanalgesia (Rompun, 0015 ml kg-' BW; Bayer, FRG) with atropine (0 04 mg kg-1 BW; Polfa, Poland). A heat exchanger was implanted on the left cervical vagus nerve; after rotation of the animal, the second heat exchanger was implanted on the right vagus, as well as a catheter to the pancreatic accessory duct and a cannula to the jejunum. The vagi were reached from two separate incisions 10 cm long (Podgurniak, 1987) in the central part of the neck, dorsal and parallel to the jugular vein. After careful and gentle isolation of the nerves the heat exchangers were implanted. The connecting tubes of the heat exchanger were exteriorized through the skin below the incision. Before suturing, the antibiotic (Penicillinum crystalisatum 300000 U, Polfa) was administered directly into the wound. Catheterization of the pancreatic duct was performed using a modification of the original method from Butler, Brinkman & Klavano (1960). A Silastic catheter (Dow-Corning, Belgium) 23 cm long and 3 x 2 mm was used. The catheter was perforated with 1 2 mm holes 25 mm from its top up to the end. Two silicon cuffs were glued onto the catheter behind the last holes to ensure proper fixation in the pancreatic duct. A cuff of 10 mm diameter mounted in the middle of the catheter immobilized it between the abdominal muscles. The catheter was introduced into the pancreatic accessory duct 5 mm from its duodenal orifice. Implantation of the jejunal perforated T-shaped (P-T) cannula was done according to Pierzynowski, Westrom, Karlsson, Swedensen & Nilsson (1988). In brief, a silicon (P-T) cannula was implanted into the jejunum, 10 cm behind the duodenal orifice of the pancreatic accessory duct, and fixed by external retaining cuffs. From 12 h after the operation and between experiments, the pancreatic catheter was connected to the jejunal P-T cannula to allow a free flow of pancreatic juice.

Experiments

Experiments with the cold blockade of the vagi started 14 days after surgery. Heat exchangers were cooled by water at 4 'C. The flow of cooling water was established at 100 ml min-'. The experiments started after 18 h fasting, with 4 x 30 min basal pancreatic juice collections. Pancreatic juice was then collected for 2 x 15 min during vagal blockade. Experiments were completed by 4 x 30 min collections of pancreatic juice. During experiments, pancreatic juice was not reintroduced into the jejunum. Samples of the pancreatic juice were placed on ice and stored frozen (-21 'C) until analysis. Analyses

Volume, total protein content (according to Lowry, Rosebrough, Farr & Randall, 1951) as well as trypsin activity (according to Erlanger, Kokowsky & Cohen, 1961) were measured in collected samples. Trypsinogen in the pancreatic juice was activated by enterokinase (Sigma) at a concentration of 1 mg ml-1 in Tris buffer, pH 7-4 at a proportion of 4:1. The units of trypsin (U) were defined as 1 the amount of enzyme which catalysed

The results were means + S.E.M.

4umol of benzylarginine-p-nitroanilid.

statistically evaluated using analysis of variance and

are represented as

RESULTS

pancreatic juice before cooling of the nerve was kg-1 vagi resulted in a decrease of pancreatic juice volume secretion to 0095 + 0-03 ml h-1 kg-' BW. After rewarming the nerves, the volume of the pancreatic juice rapidly recovered to values similar to those before cooling (0-36 + 0 07 ml h-1 kg-' BW) (Fig. 1 A). Protein concentration decreased from a basal value of 2 15 + 0-6 to 177 + 0-5 g I` during cooling and increased to 2-57 + 05 g 1-1 after rewarming the nerve (Table 1). Protein excretion fell from 0-70 + 0-15 to 0 25 + 0 03 mg h-1 kg-' BW and recovered to 0-80 + 0-16 mg h-1 kg-' BW after rewarming of the nerve (Fig. 1 B). The concentration of Mean volume 0-31 + 005 ml h-W

of

BW. Blockade of the

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COLD BLOCKADE OF VAGUS AND PANCREAS SECRETION

0.4

403

A

-~0.2

0-93

03'0 0.9

B

0-1 0.7

0-5 0

0.4

03 02 0.1

c 'O

.2

3

2

-120 -90

60-30

0 30 60 90 120 150 Time (min) Fig. 1. Volume (A), protein excretion (B) and trypsin excretion (C) in pancretic juice (bar represents period of cooling of the vagus nerve). Values represent mean ±+S.E.M.

trypsin decreased from 9 09 + 2-28 to 6-48 + 1c8 1 U ml-' during cooling and returned to the basal value (I10-2 + 1 .79 U ml-') after the blockade of the nerve (Table 2). Pancreatic trypsin excretion decreased from 3 05 + 0 69 U h-' kg-' BW in the control period to 0-84 +0-23 U h-1 kg-' BW during the cooling and returned to a basal value of 3-33 + 0 52 U h-1 kg-' BW after cooling (Fig. I C). Downloaded from Exp Physiol (ep.physoc.org) by guest on July 10, 2011

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Table 1. The concentration of pancreatic juice protein (g L-') in the samples collected before (1-4), during (5, 6) and after (7-10) cooling of the vagus nerve No.

Mean

+ S.E.M.

1 2 3 4 5 6 7 8 9 10

2-76 2-38 2-90 2-30 1-68 2-03 2-77 2-23 2-18 2-62

0-73 0-63 1-07 054 0 49 0 45 0-32 0 51 0-42 0 65

Table 2. The concentration of trypsin (U ml-1) in the pancreatic juice before (1-4), during (5, 6) and after (7-10) cooling of the vagus nerve No.

Mean

+ S.E.M.

1 2 3 4 5 6 7 8 9 10

10-7 85 10-1 90 8-2 6-7* 10-8 94 10-7

2-7 2-2 2-9 2-2 3-3 1-6 0-8

11-2

2-1 2-7 2-8

* Statistically significant (P < 0 05).

DISCUSSION

The blocking action of cold on nerve conduction has been described by Boyd & Ets (1934), Douglas & Malcolm (1955) and Paintal (1965). This effect was exploited to investigate the effects of interrupting the vagal supply on respiration, on the transmission of impulses by atrial receptors (Linden, Mary & Weatherill, 1981), on gastrointestinal motility (Hall, ElSharkawy & Diamant, 1982) and on the pancreatic secretion (Magee et al. 1965; Lenninger, Magee & White, 1965; Grundy et al. 1983). Observations recently made on conscious sheep have confirmed many of these findings (Podgurniak, 1987, 1988). In the experiments reported here, cold block of the vagi of conscious calves resulted in a decrease in volume (29 %), protein content (36%) and trypsin activity (27 %) of the pancreatic juice. Rewarming restored all these variables to their initial values. The effect of cold block was immediate. There was no increase in the secretion rate above control levels after rewarming in contrast to the findings of Grundy et al. (1983). However, in the latter experiments, pancreatic secretion was stimulated continuously by pure secretin, even during the period of cold block, so the experiments are not directly comparable. These experiments provide further evidence that the vagus provides a drive to pancreatic Downloaded from Exp Physiol (ep.physoc.org) by guest on July 10, 2011

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secretion, either through vago-vagal reflexes or through some central mechanism and that in the ruminant fasted 24 h, the vagus must contribute about one-third of the total pancreatic response. REFERENCES

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