Functional nasal salt glands in adrenalectomized domestic ducks (Anas platyrhynchos)

GENERAL

AND

COMPARATIVE

Functional

ENDOCRINOLOGY

4,

15-26 (1980)

Nasal Salt Glands in Adrenalectomized

Domestic

Ducks

(Anas p/a tyrhynchos) DAVID Department

of Zoology,

University

GORDON

BUTLER

of Toronto.

Toronto,

MSS

IAI.

Ontario,

Canada

Accepted July 10, 1979 Rates of fluid, Na’, and osmolyte secretion by the nasal salt glands were unchanged following surgical adrenalectomy. However, adrenalectomy did shorten the period of secretion, thereby decreasing the total output of fluid to 43 (2 days postoperatively), 72 (7 days), and 53% (14 days) of the total volumes secreted by the respective controls. Consequently there was a significant reduction in the total output of both Na’ and osmolytes. The interval between the start of intravenous loading (0.5 M NaCI) and the onset of secretion was less than 3 min in both adrenalectomized and sham-operated ducks. Adrenalectomy was verified by histological examination and measurements of plasma corticosterone. Adrinalectomized ducks adapted to 0.9410NaCl by increasing Na’ and osmotic concentrations in the nasal fluid. Spontaneous nasal secretion was observed in adrenalectomized ducks.

Adrenocortical steroids may control the rate of fluid secretion by the avian nasal salt glands (NSG) by regulating the active transport of Na+ through the secretory epithelium (Phillips et al., 1961; Fletcher et al., 1967; Holmes et al., 1970; Thomas and Phillips, 1975b). Active NSG accumulated radioactivity following the intravenous injection of [3H]corticosterone whereas inactive glands contained relatively small amounts of the isotope. Recently it has been shown that corticosterone and/or its metabolite ll-dehydrocorticosterone are bound by cytosol and nuclear proteins from duck NSG (Sandor and Fazekas, 1974; Allen, 1975; Allen et al.. 1975; Sandor et al., 1977). In general, these observations have supported the hypothesis that NSG function is steroid dependent. In an earlier experiment (Phillips et al., 1961) we studied NSG function 6-12 hr after bilateral surgical adrenalectomy. More recently, Thomas and Phillips (1975b) repeated and improved the procedure for adrenalectomy and increased the recovery period to 2 days. In the initial experiments NSG function often stopped completely whereas in the recent experiments (Thomas and Phillips, 1975b) secretion was transient.

In both experiments, severe or total impairment of NSG function may have been caused by a decrease in blood pressure and/or impaired regional blood flow to the NSG, accompanied by a renal Na+ and water diuresis in the absence of aldosterone, a naturally occurring steroid in ducks (Donaldson and Holmes, 1965; Thomas and Phillips, 1975a). The present report is based on a study of NSG function in adrenalectomized domestic ducks that have been allowed as much as 14 days to recover after surgery. It considers the possibility that the severe impairment of NSG function may have been due largely to dehydration and/or to cardiovascular failure and not, as originally proposed (Phillips et al., l%l), to the effect of adrenal insufficiency on the cellular transport of Na+ by the salt glands. MATERIALS

AND METHODS

Male white Pekin ducks (Anas platyrhynpurchased from King Cole Ducks Ltd., Aurora, Ontario, and held under a normal (12L:l2D) photoperiod in the Department of Zoology. Their regular diet, both before and after surgery, consisted of tap water and commercial duck grower food ad libitrrm. The ducks were about 7 weeks old at the time they were used for the experiments. They weighed about 3.0 kg. Animals. chos) were

15 0016-6480/80/010015-12$01.00/O Copyright 0 1980 by Academic Press, Inc. All rights of reproduction in any form reserved.

16

DAVID

GORDON

BUTLER

E.uperimentul groups. A group of live ducks was nares with a plastic nose cone as described by Phillips unilaterally adrenalectomized and allowed I week to ct al. (1961). The cotton was removed at 5-min interrecover before the second adrenal was removed. vals and reweighed to determine the fluid volume by These totally adrenalectomized ducks were then fed difference. This procedure was repeated throughout 0.9% NaCl instead of tap water. NSG function was the entire course of nasal salt gland secretion. The assessed 2. 7, and 14 days later or as long as the ad- cotton samples were dried in cacao and the solute was renalectomized ducks survived. A second group of redissolved in 10 ml of distilled water for Na’ analysis five ducks was sham-operated and used as controls so by flame photometry (Zeiss Model PF5). The osmotic that the effects of surgical trauma and/or handling concentration of these samples was determined by could be accounted for. freezing point depression with a Fiske osmometer. Proc~edare ,fiw adrenalectomy. The right wing was Sodium and osmotic concentrations were also deteranesthetized with lidocaine (Xylocaine-Astra) and an mined in plasma samples. Plasma glucose was measured by the glucose oxidase method (Worthington indwelling cannula was inserted into the brachial vein. Approximately 3.0 ml/kg body wt of Equithesin (100 Diagnostics). ml aqueous solution containing Na-pentobarbitol, 0.96 Impro\~ement of the method for adrrnalectomy. g: MgSO,, 2.12 g; chloral hydrate. 4.26 g; ethanol, 8.0 Before the major part of the experiment was started, a ml; propylene glycol. 20.0 ml-see DesForges and significant number of ducks were adrenalectomized for Scott, 1971) was injected into the cannula. When the practice. These ducks were fed tap water both before duck had become unconscious it was tied to a holding and after surgery. On tap water, they soon became board with the left side uppermost. The feathers were weak, dehydrated, and they generally died within 4 to removed from the surgical area and the skin was 5 days after the second adrenal was removed. Some of washed with 7% ethanol. An incision was made on these adrenalectomized ducks were examined to show the left-hand side between the vertebral segments of whether or not partial or total adrenalectomy had the fifth and sixth ribs. The adrenal was removed using modified the response of the nasal salt glands to intrathe procedure described by Thomas and Phillips venous loads of 45 ml of 0.5 M NaCI. (1975a). Access to the adrenal was considerably easier Completeness of adrenalrctomy. At autopsy, areas if the left testicle was removed first. With practice it in which the adrenals had been located were carefully was possible to remove the adrenal intact with the loss examined under a Zeiss operating microscope. All of of less than 3 ml of blood. The area was packed with the surrounding tissue was excised, fixed in Bouin’s Gelfoam sponge (Upjohn) and the ribs were pulled to- solution, sectioned (10 pm), and stained with gether with stainless-steel wire. The wound was closed hematoxylin and eosin. These sections were carefully with Michel clips and bandaged. The duck usually re- examined for residual adrenocortical tissue in order to gained consciousness within 30-60 min after the verify completeness of adrenalectomy. surgery was completed. The operated ducks were held Radioimmanoassay for cwticosterone. Corticostein isolation cages overnight and returned to the flock rone levels in the control plasmas (ducks acclimated to the following day. One week was allowed for recovery freshwater or 0.9% saline) and in those collected from before the right adrenal was removed. On the right experimental birds 7 days after bilateral surgical adside, entry was made between the vertebral segments renalectomy were determined using the method deof the sixth and seventh ribs following cannulation of veloped for poultry by Etches (1976). Plasma samples the left brachial vein. The right testicle was not re- collected from the same ducks I4 days after admoved. Both sham-operated and adrenalectomized renalectomy were assayed using the following general ducks were injected regularly with penicillin G procedure. Duplicate IO-PI samples of plasma were (Pfizer). Any operated ducks that showed signs of in- diluted with 500 ~1 of assay buffer, and the solution fection or weakness were not used for experimental heated to 98” for 10 min to destroy the corticosteroid assessment of NSG function. binding globulins. Anti-corticosterone (CO 21) and Procedure for collecting nasal j7aid samples. Each [3H]corticosterone were then added and the samples test duck was tied loosely to a holding board. After a were incubated overnight at 4”. Free corticostefew minutes the bird settled into position and there rone was adsorbed onto dextran-coated charcoal. Perwas no evidence of stress. A preload blood sample (3 centage bound [3H]corticosterone was plotted against ml) was withdrawn from the left wing-vein cannula, corticosterone standards and the corticosterone conthen 45 ml of 0.5 M NaCl was slowly infused into the centration of the unknowns were derived from the right brachial vein cannula. At the first sign of secre- standard curve. Using this procedure, the lower limit tion from the NSG another 3-ml blood sample was of detection for cotticosterone was 0.10 &IO0 ml withdrawn from the left brachial vein in order to de- plasma; for Etches’ procedure, 0.03 cLg/lOOml plasma. termine the Na and osmotic concentrations at the Statistics. Mean values for sham-operated and adonset of secretion. renalectomized data were compared using a single Nasal fluid was collected onto preweighed, washed classification ANOVA (Model I) for two groups with cotton which was held in position over the external equal and unequal sample sizes. Changes in mean val-

Oct. 26

Dec. I Dec. 2 Dec. 6

Jan. 5

Jan. 18

Jan. 19

Feb. 8

87

30 33 39

69

34

35

48

2

3

2

2

30 13 4

Intact

TABLE

1

No secretion

No secretion

No secretion

No secretion

12 15 3 after second load

2

Time to onset of secretion (min)

-

-

-

-

17.4 22.0 3.1

26.4

Total volume (ml)

ON NASAL SALT GLAND FUNCTION BEFORE AND AFTERSURGERY"

45 ml of 0.5 M NaCl Same Same 2 x 45 ml of 0.5 M NaCl with I-hr interval 45 ml of 0.5 M NaCl 2 x 45 ml of 0.5 M NaCl with 65-min interval 2 x 45 ml of 0.5 M NaCl with 67-min interval 45 ml of 0.5 M NaCl

Intravenous load

ADRENALECTOMY

IN

-

-

-

-

504 + 12 501 2 4 455 2 20

421 +4

Na+ (meqiliter)

-

-

-

-

683 ” 39 643 iz 38 932 ic 36

828 ?z 6

mOsm/liter

None

None

None

2

Intact adrenals 55 30 11

Adrenal remnants (mg)

DOMESTIC DUCKS FED WITH TAP WATER

UValues are means i- SE. Saline was injected into the right brachial vein. Duck No. 87, an intact control, is included for comparison.

Date

SURGICAL

Days after adrenalectomy

OF BILATERAL

Duck No.

EFFECT

5

z E m

9 9 2 !z K r: 2 z E E

:! P E w

r;

ic

DAVID

18

GORDON

ues within groups following various treatments were assessed using a f test for paired comparisons.

RESULTS

Table 1 shows that the NSG were functional in ducks with 11, 30, and 55 mg of adrenal remnants. In four ducks with 2 mg of residual adrenal or less, the NSG were inactive, there being no response to one or more intravenous loads of 45 ml of 0.5 M NaCl. These data show that adrenalectomy, using this method, adequately prevented NSG secretion in ducks maintained on fresh water. The total output of fluid decreased as a function of the decreasing amounts of residual adrenal tissue. There was no concurrent trend in Na+ or osmotic concentrations in the nasal fluid. They remained fairly constant. Survival of the bilaterally adrenalectomized ducks was prolonged if they were fed 0.9% saline instead of tap water. Four

BUTLER

of a total of five adrenalectomized ducks were still alive 14 days after surgery. Nasal fluid samples were collected from only two ducks. The third secreted a trace of nasal fluid and was generally weak; the brachial vein cannulae became blocked in the fourth duck so that it was impossible to collect nasal fluid samples in the regular way. A thorough microscopic examination at autopsy failed to reveal any adrenal remnants. There was no evidence of adrenal tissue in the lo-pm serial sections through the tissue in which the glands had been embedded. Table 2 shows that plasma samples from sham-adrenalectomized ducks contained corticosterone but there was no evidence of significant amounts of corticosterone in plasma samples from bilaterally adrenalectomized ducks. The interval between saline loading (0.5 M NaCl) and the onset of nasal secretion

TABLE 2 CORTICOSTERONE IN PLASMA FROM BILATERALLYADRENALECTOMIZEDDUCKSWITH FUNCTIONAL NASALSALTGLANDS" Plasma corticosterone Intact freshwater Intact freshwater Intact (0.9% NaCl) Duck No. Sham adrenalectomy 92 23 99 88 33 Mean Adrenalectomy 40 80 52 4 24 Mean

(~gi100 ml)

1.00 0.96 0.67 7 days 0.13 (2) 0.50 1.45 0.72 0.74 0.71 + 0.22 0.04

0.07 0.07 0.06 0.04 0.05 2 0.01

14 days 1.20 0.40 2.70 0.60 2.00 1.38 -t 0.43 Not Not Not Not Not

detectable (7) detectable detectable detectable detectable (7)

n Values are means + SE. Corticosterone was measured in plasma samples collected 7 days after sham-adrenalectomy or adrenalectomy using the method of Etches (1975). This method was also used for plasma from the intact ducks. The Wday plasma samples were assayed using a different radioimmunoassay (see Materials and Methods). Numbers in parentheses indicate number of days after surgery when different from the category. Note that these operated ducks are the ones for which data are given in Tables 3-7.

SALT

GLANDS

IN ADRENALECTOMIZED

was less than 3 min in both sham-operated and adrenalectomized ducks, there being no evidence that the interval had increased following adrenalectomy. Table 3 shows that neither plasma Na+ nor osmotic concentrations changed following .adrenalectomy. Any changes that might otherwise have been evident may have been overshadowed as a result of feeding the ducks 0.9% saline. Plasma Na+ concentrations in preload samples (Table 3) were about lo-15 meq/liter higher than in ducks maintained on tap water (Holmes et al., 1961) although plasma osmotic concentrations were within the range for freshwater ducks (Stewart, 1972). Two, seven, and fourteen days after surgery the increase in osmolarity at onset of secretion was approximately 6% in both adrenalectomized and sham-operated ducks (Table 3). Plasma Na+ concentrations followed the same pattern. Table 4 shows that the rate of nasal fluid secretion (5min intervals) in adrenalectomized ducks was not significantly different

from the rate in sham-operated controls in any of the three successive collection periods (2, 7, or 14 days postoperatively). There was, however, a progressive and statistically significant increase in 5-min flow rates in both sham-operated and adrenalectomized ducks as they gradually adapted to 0.9% saline (Table 4). The flow rates had almost doubled in the 7- and 14day collections as compared with the 2-day collections. Although nasal fluid Na+ concentrations were unaffected by adrenalectomy they tended to increase in response to 0.9% saline. Together, the increased flow rates and increased Na’ and osmotic concentrations resulted in a significant increase in the rate of secretion of Na+ and osmolytes. Even though adrenalectomy had no measurable effect on the rates of fluid and osmolyte secretion by the nasal salt glands, it significantly reduced the time span of functional activity 2 days after surgery (Table 5). Seven days after adrenalectomy it was difficult to distinguish between the two

TABLE Na+ AND

PLASMA

OSMOTIC

CONCENTRATIONS AND AFTER THE

3

IN ADRENALECTOMIZED INTRAVENOUS INJECTION

AND OF

[Na’] Days after surgery

19

DUCKS

BEFORE

Osmolarity

Before

5

153.0 t2.0

289.1 k4.5

161.0 21.0

316.0 k-4.5

156.1 kO.5

299.3 23.5

5

154.4 ~1.6

289.8 ~~5.6

157.9 ~2.5

297.2 +2.8

157.4 23.1

292.6 k5.9

5

156.0 21.5

295.2 23.9

165.0 to.9

314.8 23.5

153.0 22.2

301.1 ~3.2

ADX

4

152.3 22.8

286.4 k5.9

157.5 t4.0

300.0 ~4.2

154.5 22.5

2%.6 24.4

Sham ADX

5

155.4 21.8

296.5 +3.1

162.1 io.9

313.6 k2.7

154.2 21.1

326.5 i. 18.4

2

155.6

290.5

156.5

302.6

154.8

301.4

ADX

Onset

DUCKS

NaCP

n

Sham

loading

SHAM-OPERATED

0.5 M

of secretion

End of secretion

2 ADX

Sham

ADX 7

14 ADX ’ Values

are means

+ SE (except

for 1Cday

ADX);

[Na+]

in meq/liter;

osmolarity

(mOsm/liter).

5

Sham ADX

2

0.47 20.06

492.9 t 13.8

509.7 -c 13.0

515.9 -+4 .9*:v

493.3 k-7.4”

464.8 k4.5

469.4 26.6

Na’ concentration (meq/liter)

227.2 +23.7

312.2 +32.3

270.6 280.2

239.2 +22.7

119.5 t8.0

157.1 k 12.4

Rate of Na’ (meq/kg/S min)

776.3 k42.6

364.4 k71.7

515.6 274.8

E ? ;

5 136.0 233.0 899.9 250.0

8 E s

6 452.8 +39.6 417.3

223.5 -c 15.1

257.9 218.2

Rate of osmolyte excretion (pOsmlkgi5 min)

883.1 234.0 776.7

833.4 +61.0

751.2 522.4

Osmolarity (mOsm/liter)

” Nasal fluid samples for each duck were collected every 5 min during the period of secretion. Average values (n 1 15) for each parameter were used to calculate the means % SE recorded in this table. i/zP CC0.01 compared with t-day Sham ADX. w P < 0.01 compared with 2-day ADX.

14

0.59 20.05

5

Sham ADX

ADX

0.53 +O.lS

4

0.48 kO.05

0.28 +0.004

0.33 20.03

tlow rate (ml/kg/S min)

ADX

7

5

2

5

n

ADX

Sham ADX

Days after surgery

TABLE 4 RATES OF FLUID, Na’. AND OSMOLYTE SECRETION AND NASAL FLUID Na’ AND OSMOTIC CONCENTRATIONS IN DOMESTIC DUCKS FOLLOWING BILATERAL SURGICAL ADRENALECTOMY”

SALT

TABLE PERIOD OF NASAL IN SHAM-OPERATED DUCKS

FED

ON

SALT

0.9%

GLANDS

IN ADRENALECTOMIZED

5

GLAND SECRETION AND ADRENALECTOMIZED

SALINE Days

tomy the volume was only 43% of that for the controls although 7 days after surgery it was 72% of the control volume, largely because the volume secreted by the shamoperated ducks had decreased (Table 6). The total output of both Na+ and osmolytes was significantly reduced following adrenalectomy, primarily as a result of the lowered total output of fluid (Table 6). The nasal salt glands in both shamoperated and adrenalectomized ducks responded to 0.9% saline through an increased rate of fluid and osmolyte secretion but surprisingly, the period of secretion tended to decrease (Table 5). Table 7 shows that sham-adrenalectomy was not followed by a significant change in plasma glucose concentrations when compared with intact controls. After unilateral adrenalectomy plasma glucose tended to increase whereas 2 days after bilateral adrenalectomy there was a reversal and plasma glucose decreased by 15% compared with the sham-operated ducks. This

(MIN)

POSTOPERATIVELY” after

surgery

NO.

2

I

14

Sham ADX 99 23 92 88 33 Mean

125 145 160 220 140 158+ 17

95 55 90 120 105 93k 11

95 40 15 95 120 85 -t 13

ADX 4 80 52 49 24 Mean

45 90 120 80 70 81 k 12

Died 85 70 loo 53 77 f 10

Dislodged

cannula 80 Transient secretion 53 67

” Nasal secretion was induced by an intravenous ml of 0.5 M NaCl. Values are means -t SE.

load of 45

groups. This shortened period of secretion was, in turn, reflected in a lowered total output of nasal fluid from the adrenalectomized birds. Two days after adrenalecTABLE TOTAL

OUTPUT

OF FLUID,

Na’,

AND

BILATERAL

Days after surgery Sham ADX

21

DUCKS

6

OSMOLYTES SURGICAL

BY DUCK

NASAL

SALT

GLANDS

AFTER

ADRENALECTOMY”

Total osmolytes n

Total volume (ml)

Total Na+ (meq)

(mOsm)

5

36.8 54.4

17.68 51.71

27.47 22.72

ADX

5

15.8* t 1.7*

6.96 %0.91%

12.99 a2.22”

Sham ADX

5

31.1 23.0

16.30 22.00

26.89 r1.99

ADX

4

22.5 *3.7***

11.60 + 1.94***

17.64 - .48** +3

Sham ADX

5

30.3 +4.1

14.68 e1.55

26.15 r2.66

2

16.0

8.13

12.27

2

7

14 ADX 0 Values

are means

*P < 0.005. 231: p < 0.05. *** P < 0.20.

+ SE. (except

for

14-day

ADX).

ADX

compared

with

Sham ADX.

176.6 23.3 197.3

es.8

185.8 +5.3 179.8

25.5

TABLE

168.2 23.8 149.3 210.1

18.6

Onset

2

DUCKS

180.4 t4.8 151.8*.”

Preload

(mg%,) IN DOMESTIC

7

2 14.2

184.4 25.7 162.8

End

FOLLOWING

159.0”,” k2.9

184.4 k5.2

Preload

148.0 +6.2

171.5 24.5

Onset

7

157.0 (n = 2)

177.0 28.3

Preload

185.2 15.9

End

146.0 176.5 (n = 1) (n = 2)

172.6 k5.3

Onset

14

ducks unless shown otherwise. Preload.

183.2 t5.7

186.0 25.2

End

ADRENALECTOMY"

Days after bilateral ADX or Sham ADX

SURGICAL

I’ Values are mean glucose concentrations ? SE. n = 5 for sham-operated controls; n = 4 for adrenalectomized before saline loading: onset, when nasal secretion started: end, when nasal secretion stopped. 6 P < 0.025 compared with 2-day sham ADX. CP < 0.01 compared with 7-day sham ADX ” P < 0.10 compared ivith the “intact” value for the same group. CP < 0.025 compared with the “intact” value for the same group.

ADX

Sham ADX

Intact

GLUCOSE

7 Days after unilateral ADX or Sham ADX

PLASMA

z ;! E

H

G n g

SALT

GLANDS

hypoglycemia continued throughout and 1Cday collections (Table 6).

IN

ADRENALECTOMIZED

the 7-

DISCUSSION

During preliminary experiments, at a time when the method for adrenalectomy was being improved, bilaterally operated ducks were maintained on tap water both before and after surgery. Table 1 shows that, as the completeness of adrenalectomy increased, the total output of nasal fluid decreased. Nat and osmotic concentrations did not vary greatly, the primary adjustments in salt secretion being achieved through volume control. In fact the degree of variability in Na+ and osmotic concentrations between ducks (Table 1) can often be found in nasal fluid samples from a group of intact freshwater ducks selected at random. In one duck with 2 mg of residual adrenal tissue and in three in which removal of the glands was verified as complete, the NSG were totally inactive following one or two intravenous injections of 45 ml of 0.5 M NaCl. Once it could be shown that total adrenalectomy had prevented any secretion from the NSG it was possible to go ahead with experiments that would ultimately show that the NSG in adrenalectomized ducks remained active provided they were fed with 0.9% NaCl instead of tap water. In these saline-fed adrenalectomized ducks with functional NSG, adrenalectomy was verified histologically and confirmed by measuring plasma corticosterone concentrations (Table 2). Adrenalectomy had no measurable effect on the rate of fluid secretion or the rate of Na’ secretion by NSG in ducks which were fed 0.9% NaCl. Nevertheless, the time course of secretion was reduced following adrenalectomy so that the total output of fluid decreased and, accordingly, there was a significant decrease in the total output of both Na’ and osmolytes (Table 6) in response to an intravenous saline load. This reduction was maximal 2 days after bilat-

DUCKS

23

eral adrenalectomy whereas 7 days after surgery the adrenalectomized ducks secreted about 70% of the total fluid and osmolytes secreted by the sham-operated controls (Table 6). Both sham-operated and adrenalectomized ducks were able to adapt to saline by increasing the nasal fluid Na+ and osmotic concentrations during the interval between the second and seventh day after surgery (Table 4). The rate of nasal fluid secretion also increased significantly during this period of adaptation to 0.9% saline (Table 4). A similar adaptive response to saline by the NSG of domestic ducks has been observed by Schmidt-Nielsen and Kim (1964) who found that if ducks were reared on a combination of 1,2, and 3% salt solutions instead of fresh water, the nasal fluid Na+ concentration increased from 435 + 14 to 525 + 10 meq/liter. Fletcher et al. (1967) reported that a 30-day exposure of white Pekin ducks to drinking water containing 284 mM Na+ and 6.0 mM K+ was followed by an increase in the nasal fluid Na+ concentration from 466 * 17 to 592 ? 8.0 meq/liter. This adaptive change was accompanied by a substantial increase in the rate of weight-specific Na+ secretion and an increase in salt gland ATPase activity. The present data show that not only did the duck NSG become more active in response to 0.9% saline but adrenalectomy did not prevent this adaptive change. The present observation that the NSG are still functional in adrenalectomized ducks is in disagreement with our earlier finding (Phillips et al., 1961). In our original experiments, nasal salt gland function was assessed between 6 and 12 hr after bilateral adrenalectomy. Moreover, a significant volume of blood was often lost during the procedure. Postoperative shock together with a decrease in blood pressure and blood flow rate to the nasal salt glands may have been largely responsible for the loss of function. Recently, Thomas and Phillips (1975a) improved the method for surgical

24

DAVID

GORDON

adrenalectomy of ducks and extended the recovery period to 2 days. Even with these improvements, the total volume of fluid secreted in response to an intravenous injection of 10% NaCl(24 meq/kg body wt for 2 hr) was still only 13% of that secreted by the NSG of the sham-operated controls. Function was partially restored (38% of control volume) following an intravenous injection of 200 pg of corticosterone. In the present experiments saline-maintained adrenalectomized ducks secreted only 43% of the volume produced by the sham-operated controls 2 days after adrenalectomy but without steroid replacement. Within a week, the total volume of nasal fluid had increased to 73% of that secreted by the control ducks (Table 5). In contrast, when adrenalectomized ducks were fed tap water postoperatively (Table 1) the nasal salt glands were completely unresponsive to the intravenous saline load. Another important adrenocortical steroid in birds, 18-OH corticosterone (500 pg iv), increased the secreted volume of nasal fluid in saline-loaded domestic ducks from 13.9 2 0.6 to 22.7 _t 1.4 ml but there was no change in Na+ concentration (Lanthier and Sandor, 1973). Again, in our earlier experiments, the primary change that followed the injection of adrenocortical steroid was the change in volume secreted (Holmes et al., 1961). Cortisol, 1l-deoxycorticosterone, and aldosterone increased the output of nasal fluid by 104, 89, and 184%, respectively, whereas there was a 10% decrease in the Na+ concentration which was common to all three groups (Holmes et al., 1961). Table 4 shows that adrenalectomy had no measurable effect on the rate of secretion of fluid, Na+, or total osmolytes but did shorten the time course of secretion. Moreover, adrenalectomy did not impair the adaptive increases in flow rate and Na+ and osmotic concentrations which were observed after the ducks had been fed 0.9% saline for 1 week (Table 4).

BUTLER

In both sham-operated and adrenalectomized ducks, nasal fluid secretion began less than 3 min after the intravenous injection of 0.5 M NaCl was started. Usually, from 35-40 ml of the total 45-ml load had been delivered before the first drops of fluid appeared at the external nares. This observation differs from the earlier finding (Phillips et al.. 1961) that unilateral adrenalectomy increased the interval between stomach loading and the onset of secretion from 53 to 117 min. After bilateral adrenalectomy it was extended to 270 min. When the adrenalectomized ducks were injected with cortisol the interval was shortened to 30 min. It is difficult to assess these data since the delay in onset of secretion may have been due indirectly to impaired gastrointestinal transport of NaCl and water which had been delivered by stomach tube. A marked reduction of intestinal transport of water and electrolytes is fairly common in animals with adrenal insufficiency (Swingle and Swingle, 1965). As expected, a shorter interval between loading and the onset of nasal secretion was observed when NaCl was delivered via the intravenous route (Thomas and Phillips, 1975b). Adrenalectomy increased the secretion latency from 16 to 46 min but corticosterone replacement failed to shorten it. Table 3 shows that there were no statistically significant changes in either plasma Na+ or osmotic concentrations following adrenalectomy if the ducks were given 0.9% NaCl to drink instead of tap water. If adrenalectomized ducks were fed tap water (Thomas and Phillips, 1975a) postoperatively there was a slight, transient decrease in plasma Na+ concentrations. Table 3 shows that, overall, the onset of secretion in both sham-operated and adrenalectomized ducks started when the plasma Na+ concentration had reached 160 meq/ liter. In our earlier experiments (Phillips or al.. 1961) secretion started in shamoperated ducks when the Na’ concentra-

SALT

GLANDS

IN ADRENALECTOMIZED

tion had reached 160.6 & 2.5 meq/liter but it had reached 170.2 + 3.4 meq/liter before secretion was observed in unilaterally adrenalectomized ducks and 176.0 in the only bilaterally adrenalectomized duck which secreted. Plasma glucose concentrations increased slightly following unilateral adrenalectomy (Table 7) but there was, nevertheless, a substantial reduction in “preload” glucose concentrations following bilateral adrenalectomy. It is doubtful if this modest hypoglycemia, a change that has already been observed in other adrenalectomized birds (Veiga et al., 1978) impaired NSG function. At these concentrations, even though reduced, there would be an adequate supply of glucose to meet any metabolic needs of fully functional NSG (see Peaker et al., 1970). Adrenalectomized ducks urinated continually and at a higher rate than shamadrenalectomized ducks which were housed in adjacent cages. It was intended to counterbalance the progressive urinary natriuresis by feeding these adrenalectomized ducks 0.9% saline, thereby keeping them in reasonably good health and permitting an extended period of observation. Spontaneous NSG secretion was observed in adrenalectomized ducks. If, on the other hand, the adrenalectomized ducks were fed tap water they often became dehydrated and weak and died from 4-5 days after surgery. It is possible, although not verified by the present experiments, that the dietary intake of NaCl and water tended to offset the extensive renal Na+ and water diuresis thereby preventing dehydration and temporarily maintaining the ionic composition of tissue fluids within functional limits. It is doubtful, however, that the loss of myocardial performance, decrease in blood pressure and volume, and changes in peripheral vascular resistance which are commonly observed in adrenalectomized mammals would be repaired with 0.9%

DUCKS

25

saline but without steroid replacement (Reidenberg et al., 1963; Swingle and Swingle, 1965; Webb et a/., 1965; Lefer et al., 1968). It has already been shown that dehydration will impair nasal salt gland function in gulls. Douglas and Neely (1969) found that if herring gulls (Lams argentatus) were deprived of water for periods of up to 50 hr, there was severe impairment of nasal salt gland function in response to a 20 mM/kg load of NaCl by stomach tube. The degree of impairment of function was correlated with the amount of dehydration. Phenoxybenzamine increased nasal fluid secretion in these dehydrated gulls, from 20-30% of the rate in intact control ducks to 65-70%. It was argued that phenoxybenzamine’, an c-u-adrenergic blocking agent, had prevented the constriction of blood vessels supplying the salt glands. In the present experiments, the primary change after adrenalectomy was the shortened period of secretion and, as a result, a significant reduction in the total volume of fluid secreted by the salt glands. Whether or not this change was due to dehydration and/or cardiovascular failure which are characteristic changes in adrenalectomized mammals (Reidenberg et al., 1963; Webb et al., 1965: Lefer et al., 1968) remains to be shown. ACKNOWLEDGMENTS The author gratefully acknowledges continued support from the National Research Council of Canada (Grant A23.59) and technical assistance from Ms. Winnie Leong. Dr. T. K. Bhattacharyya cut and examined sections of tissue. Mr. A. Gureky, Radioassay Systems Laboratories, Carson, California. and Dr. R. S. Etches and Ms. C. Duke. Department of Animal and Poultry Science. University of Guelph kindly performed the radioimmunoassays for plasma corticosterone.

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