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Department of Surgery, Brown University School of Medicine, and the Providence Veterans Affairs Medical Center, Providence, Rhode Island 02908
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ABSTRACT |
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 TOP
 ABSTRACT
 INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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In this study, we
investigated the way in which fetal insulin secretion is influenced by
interrelated changes in blood glucose and sympathoadrenal activity.
Experiments were conducted in late gestation sheep fetuses prepared
with chronic peripheral and adrenal catheters. The fetus mounted a
brisk insulin response to hyperglycemia but with only a minimal change
in the glucose-to-insulin ratio, indicating a tight coupling between
insulin secretion and plasma glucose. In well-oxygenated fetuses,
2-adrenergic blockade by idazoxan effected no change in
fetal insulin concentration, indicating the absence of a resting
sympathetic inhibitory tone for insulin secretion. With hypoxia, fetal
norepinephrine (NE) and epinephrine secretion and plasma NE increased
markedly; fetal insulin secretion decreased strikingly with the degree
of change related to extant plasma glucose concentration. Idazoxan
blocked this effect showing the hypoxic inhibition of insulin secretion
to be mediated by a specific 2-adrenergic mechanism.
2-Blockade in the presence of sympathetic activation
secondary to hypoxic stress also revealed the presence of a potent
-adrenergic stimulatory effect for insulin secretion. However, based
on an analysis of data at the completion of the study, this
-stimulatory mechanism was seen to be absent in all six fetuses that
had been subjected to a prior experimentally induced hypoxic stress but
in only one of nine fetuses not subjected to this perturbation. We
speculate that severe hypoxic stress in the fetus may, at least in the
short term, have a residual effect in suppressing the -adrenergic
stimulatory mechanism for insulin secretion.
hyperglycemia; catecholamines; idazoxan; propranolol; hypoxia
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INTRODUCTION |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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INVESTIGATION OF THE
CONTROL of fetal insulin secretion has consisted largely of in
vitro rat studies, a modest number of studies in subhuman primates and
humans, and in vivo studies in sheep. In vitro studies in the rat have
been concerned with the ontogeny of the quantitative insulin response
to various secretagogues and specific mechanisms of stimulus-secretion
coupling in control of insulin secretion (20, 24, 32). Of
particular interest to our work, adrenergic inhibition of insulin
secretion was demonstrated in late gestation fetal rats and in fetal
rat pancreatic -cells (16, 22). In monkeys a
minimal fetal insulin response to glucose was found in acute
experiments, presumably with sympathoadrenal activation and adrenergic
inhibition of insulin secretion (30); we subsequently
noted a brisk insulin response to glucose in a more physiological
chronic fetal monkey preparation (10). An increasing fetal
insulin concentration and insulin-to-glucose ratio (I/G) were found
from 17 to 38 wk in human pregnancies (12). In this same
study, a lower I/G was seen in smaller fetuses suggesting "pancreatic -cell dysfunction."
Investigations of insulin secretion in fetal sheep have been conducted
for the most part in chronic in vivo preparations and most often have
been concerned with the insulin response to secretagogues, especially
glucose (2, 5, 6, 29). Studies of the neurohumoral control
of insulin secretion in the sheep fetus demonstrated inhibition by
epinephrine (Epi) of both basal and glucose-stimulated insulin secretion (14) with the speculation advanced that the
previously reported suppression of insulin by fetal hypoxia
(34) was secondary to associated increases in
sympathoadrenal activity. We confirmed in fetal sheep the
inhibitory effect of hypoxemia on the insulin response to hyperglycemia
and found a concurrent increase in plasma norepinephrine (NE) and in
adrenal secretion of NE and Epi (25). In that same work,
using the -adrenergic blocker phentolamine, we demonstrated this
inhibition to be -adrenergic in type. However, as phentolamine is a
general -adrenergic blocker, the specific nature of the
-inhibitory mechanism involved was not defined. Moreover,
conclusions were confounded by the fact that phentolamine directly
stimulates insulin secretion (33).
The present work was designed to test the following hypotheses:
1) in the fetus, insulin secretion is closely matched to
extant glucose concentration, 2) resting sympathetic
inhibitory tone for insulin secretion is absent in the well-oxygenated
fetus, 3) stress-initiated sympathetic activation partially
inhibits insulin secretion in the fetus, and 4) this
sympathetic inhibition of insulin secretion is mediated by a
specific 2-adrenergic mechanism.
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METHODS |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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Experimental preparation. Experiments were conducted in mixed-breed ewes with dated singleton pregnancies. Gestational age at the time of the preparative procedure ranged from 119 to 123 days, and at parturition it ranged from 142 to 147 days.
Fetuses were prepared with chronic peripheral and adrenal cannulas as described previously (9). In brief, under general anesthesia, a femoral arterial catheter was placed in the ewe; femoral arterial and venous catheters were placed in the fetus. A fetal adrenal cannula was established by insertion of a polyvinyl catheter distally in the left renal vein, excision of the left kidney, and placement of a hydraulically activated choker around the renal vein close to its junction with the vena cava. With the choker open and the catheter occluded, adrenal effluent followed its normal path via adrenal veins into the renal vein and then into the vena cava. With the catheter open and the renal vein occluded by the choker, adrenal outflow was diverted into the catheter. A minimum of 7 days was allowed for recovery from surgery before experiments were performed. A companion ewe, housed with the experimental animal, was present in the laboratory during all studies. Sample collections were begun a minimum of 60 min after the animals were moved into the laboratory. Maternal inspired O2 fraction (MFIO2) was adjusted by the insufflation of either 100% O2 or 10% O2 plus 2.5% CO2, balance nitrogen, into a plastic bag placed over the head of the ewe.Biochemical methods. Arterial gases were measured by a blood gas analyzer (model 168; Ciba Corning Diagnostics, Medfield, MA) at 37°C; hemoglobin and oxygen saturation were assessed on an OSM2 Hemoximeter (Radiometer, Copenhagen, Denmark); blood oxygen content was calculated from hemoglobin, PO2, and O2 saturation. Plasma glucose and lactate concentrations were determined on a YSI glucose-lactate analyzer (model 2700 Select; Yellow Springs Instrument, Yellow Springs, OH). Plasma immunoreactive insulin was analyzed by a solid phase 125I RIA kit (Diagnostics Products, Los Angeles, CA). The intra- and between-assay coefficients of variation for insulin were <5.2%.
Catecholamines in plasma were measured by HPLC with electrochemical detection (19, 21). Adrenal vein and peripheral blood samples were chilled on collection, and the plasma was separated in a refrigerated centrifuge at 4°C within 15 min. Plasma was frozen at
80°C until assay. For the catecholamine analysis, plasma samples
were mixed with the internal standard dihydroxybenzylamine. The
catecholamines were adsorbed onto 50-mg acid-washed aluminum oxide (Woelm Neutral; ICN Nutritional Biochemicals, Cleveland, OH) by
mixing with Tris buffer at pH 8.5. The alumina was washed twice with
water, and the catecholamines were eluted with 100 µl of 0.1 M
HClO4, using a centrifugal microfilter [model MF-5500, Bioanalytical Systems (BAS), West Lafayette, IN]. The resulting acidic
extract was injected into a chromatographic system (BAS 200A), which
used an amperometric detector (LC4B, BAS) with a potential of 0.065 V
vs. a silver-silver chloride electrode. A proprietary mobile phase
(CF-1100) and catecholamine column (MF-6213CL), both from BAS, were
utilized at a constant temperature of 30°C. Catecholamine
concentrations were determined by peak height determination in relation
to the internal standard. The intra- and between-assay coefficients of
variation were <10%. The limits of sensitivity were 20 pg of Epi or
NE injected onto the column. Adrenal secretion rates were calculated by
subtracting the peripheral arterial plasma concentrations of NE and Epi
from those in adrenal venous plasma and multiplying by adrenal plasma
flow (9).
Experimental design. From one to four experiments were conducted in random order in each animal with a minimum of 2 days of rest between experiments, which allowed for measured variables to return to original baseline. Studies were performed at the same clock time to avoid the possible influence of circadian variations. Fetal and maternal heart rate and blood pressure were recorded continuously, except during brief periods of peripheral sampling (transducer, model 1280B; recorder, model 7700; and rate computer, model 8812A; Hewlett-Packard, Waltham, MA). Values for fetal blood gases and lactate, NE and Epi secretion, and plasma NE, glucose, and insulin concentrations were determined at 30-min intervals. Sampling for NE and Epi secretion consisted of three consecutive precisely timed 2-min collections of adrenal venous effluent from which a mean value was derived.
Each study was 6 h in length and followed the same basic format with sampling as above during three 2-h periods or segments. Segment 1 consisted of baseline observations. In segment 2, fetal oxygenation was enhanced by a MFIO2 of 100% to ensure against fetal hypoxia. During this period either euglycemia (saline infusion) was maintained or hyperglycemia (fetal glucose infusion at 20 mg · min
1 · kg1 estimated
fetal body wt) was initiated. In segment 3 the fetal glycemic state (euglycemia or hyperglycemia) associated with
segment 2 was continued. Additionally, this period was
characterized by either continued hyperoxia
(MFIO2, 100%) or hypoxia
(MFIO2, 10%), 2-adrenergic
blockade by idazoxan (1 mg/kg iv; Sigma Chemical, St. Louis, MO)
(11) or idazoxan plus -adrenergic blockade by propranolol (0.5 mg/kg iv; Wyeth-Ayerst, Philadelphia, PA), or no
autonomic blockade. Efficacy of the propranolol block was tested by
observation of the fetal heart rate response to 5 µg isoproterenol (Sigma Chemical) before and after propranolol administration and at the
conclusion of the experiment.
The overall study included seven separate experimental groups that are
defined by the characteristics of the final 2 h of the experiment
(segment 3) as follows: group I, hyperglycemia, MFIO2 100%; group II,
euglycemia, MFIO2
100% plus idazoxan; group III, hyperglycemia,
MFIO2 100% plus idazoxan; group
IV, euglycemia, MFIO2 10%;
group V, hyperglycemia, MFIO2
10%; group VI, hyperglycemia, MFIO2 10% plus idazoxan; group
VII, hyperglycemia, MFIO2 10% plus idazoxan plus propranolol.
Statistical analysis. Data are presented as means ± SE. Comparisons of changes within each group in glucose, insulin, PO2, O2 content, pH, PCO2, and lactate, and logs of NE and Epi adrenal secretion and plasma NE were made on the means of four observations during each 2-h sampling period with repeated-measures ANOVA. When the ANOVA was positive, pair-wise comparisons were done with the Student-Newman-Keuls test. Between group differences were studied by the t-test and by the Mann-Whitney nonparametric test. Statistical programs used were Minitab, 5.1 release, (Minitab, Pennsylvania State University) and Primer of Biostatistics: The Program (McGraw-Hill, New York, NY, 1987).
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RESULTS |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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Results are reported from seven protocols designated as groups I-VII, with groups arranged in sets according to principal conceptual points in the study. A total of 54 experiments was completed in 19 fetal preparations. Fetal hemodynamic variables (data not shown) were unremarkable and were in keeping with viable, intact, responsive fetuses.
Primary data are provided in the tables. Data are shown as means of four observations made at 30-min intervals during each of three 2-h segments: baseline control, hyperoxia with euglycemia or hyperglycemia, and specific perturbation.
Group I (Tables 1 and
2) provided basic control data for the
study. With enhanced oxygenation in segment 2,
compared with segment 1, PO2 and
arterial O2 content (CaO2) increased. With glucose infusion in segment 2, glucose and insulin levels
increased. In segment 3, with continued hyperoxia and
glucose infusion (exactly as in segment 2), there were no
further changes in oxygen levels or plasma glucose or insulin.
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Data regarding the insulin response to glucose are derived from
groups I, III, V, VI, and VII, each of which
included a 2-h baseline control period (segment 1) and a 2-h
period of enhanced oxygenation and glucose infusion (segment
2). Plasma insulins for these groups are illustrated in Fig.
1. In segment 2, baseline glucose values increased about 2.5-fold, and baseline insulin values
increased by 2.5- to 3-fold. The I/G, 0.59 at baseline and 0.70 in
segment 2, were not significantly different. The I/G patterns for segments 1 and 2 are illustrated in
detail in Fig. 2, where the mean I/G is
shown for each of the four 30-min samples at baseline and for each of
the four measurements during segment 2. The I/G at 60 min of
hyperglycemia was higher than all control values, and the 90-min value
was higher than that for the 90-min control observation. There was no
difference among the hyperglycemic values.
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Groups II and III (Tables
3 and 4)
were concerned with the assessment of a resting sympathetic inhibitory
tone for insulin secretion with euglycemia and hyperglycemia. In both
groups, fetal oxygenation was enhanced and well maintained throughout
segments 2 and 3. In group II, with no
exogenous glucose infusion, plasma glucose was unchanged (from
baseline) in segment 2 and slightly decreased in
segment 3. In group III, with exogenous glucose
infusion, plasma glucose increased (from baseline) about 2.5-fold in
segments 2 and 3. There was no change in insulin
values (from segment 2) in segment 3 with
2-adrenergic block by idazoxan in the presence of either
euglycemia (Fig. 3A) or
hyperglycemia (Fig. 3B).
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Groups IV and V (Tables
5 and 6)
addressed the effect on insulin secretion of sympathetic activation by
hypoxic stress with euglycemia and hyperglycemia. In both groups, in
segment 2 with elevated
MFIO2, fetal
oxygenation was enhanced, whereas in segment 3 with
a MFIO2 of 10% there was a precipitous
fall in fetal oxygen values. During hypoxia in both groups, pH and
PCO2 decreased and lactate increased; NE and
Epi secretion and plasma NE all increased considerably. In
group IV with no exogenous glucose infusion during the
hypoxic episode in segment 3, plasma glucose increased about 30%; insulin values decreased from those in segments 1 and
2 by about 50%. In group V with exogenous
glucose infusion, plasma glucose increased (from baseline) about
2.5-fold in segments 2 and 3. Insulin values
increased almost threefold in segment 2, but during the
hypoxic episode in segment 3 decreased >50% to a value
that was not different from that at baseline with euglycemia. The I/G
during periods of hypoxic stress were not different from one another in
the presence of euglycemia (Fig.
4A) and hyperglycemia (Fig.
4B).
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Groups VI and VII (Tables
7 and 8)
were concerned with elucidation of - and -adrenergic factors in
control of insulin secretion. In both groups, in segment 2 fetal oxygenation was enhanced with elevated
MFIO2, whereas in segment 3 with MFIO2 of 10% fetal oxygen values
fell precipitously. In both groups, during hypoxia in segment
3, pH and PCO2 decreased and lactate increased; NE and Epi secretion and plasma NE all increased remarkably and to a similar degree. In both groups, with exogenous glucose infusion in segments 2 and 3, plasma glucose
increased about 2.5 times over baseline.
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In group VI insulin values increased about threefold in
segment 2; in segment 3 with hypoxic stress and
2-adrenergic blockade by idazoxan insulin levels more
than doubled compared with segment 2 (Fig.
5A). In group VII
insulin values increased about 2.5 times in segment 2; in
segment 3, with hypoxic stress and 2-block by idazoxan and -block by propranolol, insulin levels increased by only
about 25% over those in segment 2 (Fig. 5B).
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In group VI, insulin responses in segment 3 (hyperglycemia, hypoxemia, and -blockade) varied markedly among
experiments and were seen to fall into low-insulin (increase <30%)
and high-insulin (increase >90%) subgroups (Fig.
6). The percent changes in individual experiments in the low-insulin subgroup were 29, 14, 23, 26, 26, 27, and 32 (17 ± 8) and in the high-insulin subgroup were 91, 96, 135, 147, 233, 254, 269, and 283 (189 ± 28). Blood gas, glucose,
and catecholamines values were quite similar for the two subgroups
(Tables 9 and
10). I/G in segment 2 for
the low- and high-insulin subgroups (0.59 ± 0.11 and 1.11 ± 0.26, respectively) were not significantly different. However, in
segment 3 with hypoxia, the I/G for the low-insulin subgroup
(0.81 ± 0.19) was significantly lower than that for the
high-insulin subgroup (3.22 ± 1.00; P < 0.009).
The segment 3 insulin levels for the high-insulin subgroup were significantly different from those for group VII
(P < 0.009), whereas the low-insulin subgroup and
group VII were not different from one another.
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DISCUSSION |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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In this study in late gestation sheep fetuses, we examined the
response of insulin secretion to increases in plasma glucose and the
mechanisms of sympathoadrenal control of insulin release. Extensive in
vitro studies in fetal rat tissues have yielded some insight into the
ontogeny of stimulus-secretion coupling in control of insulin secretion
in that species (20, 24, 32). However, from such studies
little is known of the neurohumoral mechanisms involved other than that
Epi blocks the release of insulin by fetal rat -cells (16,
22). Studies in vivo with intact nervous connections offer
evident advantages in the investigation of this area. The sheep fetus
generally is well suited to this purpose, allowing the determination in
our experimental preparation of adrenal medullary secretion rates of NE
and Epi as well as of plasma NE. Given the critical physiological role
in sheep of catecholamines of adrenal origin in the inhibition of
insulin secretion (3) and in the fetal adaptive response
to hypoxia (8), specific inferences can be drawn regarding
the mechanisms of sympathoadrenal influences on insulin release.
Enhanced oxygenation (hyperoxia) with modest increases in fetal PO2 and CaO2 (26) was induced in segment 2 of all experiments to ensure against episodes of hypoxemia as can occur with hyperglycemia (31). This maneuver was effective in that stable fetal oxygen levels were maintained in all except one experiment in which a spontaneous transient hypoxemic episode was seen. NE secretion decreased slightly in groups I, III, and IV with hyperoxia but not in the remaining groups. The shifts in PO2 in all groups were close to the beginning inflection of the NE secretion-PO2 curve where some variability animal to animal and thus group to group could reasonably be expected (9). The absence of significant changes in Epi secretion or plasma NE indicates that physiologically meaningful changes in catecholamines did not occur. In the two groups (II and IV) in which euglycemia was maintained there were no changes in plasma glucose or insulin in segment 2 compared with segment 1. Group I served as a universal control in that experimental conditions (hyperoxia and hyperglycemia) were unchanged from segment 2 to segment 3. The absence of significant changes in glucose, insulin, or other measured parameters between these two segments indicates stability of the preparation and absence of spurious and thus confounding changes during the final 2 h (segment 3) of the experiments.
Fetal insulin response to glucose. The experimental design provided a large body of data regarding the fetal insulin response to a glucose stimulus. These data were derived from five experimental groups (I, III, V, VI, and VII), each of which included a 2-h baseline control segment and a 2-h segment of maternal oxygen breathing with fetal infusion of glucose.
The capacity of the relatively mature (>110 days of gestation) sheep fetus to mount a vigorous insulin response to elevations in plasma glucose has been clearly demonstrated (13, 23). Our study was compatible with this earlier work but yielded new data of interest. Whereas exogenous glucose infusion led to a marked increase in plasma insulin (Fig. 1), the average I/G was not significantly increased above baseline. Observation of I/G at each 30-min sampling point gives a more dynamic picture of the insulin responses (Fig. 2). The ratios at 60 and 90 min in segment 2 were modestly higher than equivalent values at the euglycemic baseline, showing a small, temporary overshoot in insulin secretion in response to exogenous glucose. Nevertheless, these overall results are remarkable for the narrow range of I/G over a wide range of glucose concentrations and show clearly a close match between insulin secretion and glucose concentration. Maintenance of stable I/G over a 2-h period suggests considerable insulin secretory capacity, although of course the metabolic clearance of insulin could also be a contributing factor in maintaining plasma insulin levels. There was also no apparent relationship between insulin or I/G and prior hypoxic stress, a point that will be alluded to below.Absence of baseline sympathetic tone inhibiting fetal insulin
secretion.
2-Adrenergic blockade with idazoxan in segment
3 induced no change from segment 2 in insulin levels in
the well-oxygenated fetus during periods of euglycemia (group
II) or hyperglycemia (group III) (Fig. 3). With the
decrease in glucose in segment 3 in group II,
there was an equivalent decrease in plasma insulin maintaining an exact
match with segment 2 in the I/G. Given the demonstrated
efficacy of idazoxan as an 2-blocking agent in
group VI, this indicates that resting sympathoadrenal tone
does not modify the secretion of insulin by the fetus in the basal
state. The marked inhibition of insulin by induced hypoxemia seen in groups IV and V (Table 5, Fig. 4) demonstrated
the potential effect of increased sympathetic activity on insulin
secretion and supports the concept that this inhibitory system is
present but inactive in the resting state. These findings are in
keeping with what has been found in adult animals (33).
The suggestion made previously that such a resting inhibitory tone for
insulin secretion does exist in fetal sheep (35) is
erroneous. That study was based on the use of the general
-adrenergic blocking agent phentolamine, which was subsequently
shown to have, in addition to its blocking action, a direct stimulatory
effect on insulin secretion (33).
Inhibition of insulin secretion by hypoxic stress in euglycemia and hyperglycemia. Hypoxic stress with its associated adrenergic activation resulted in a marked inhibition of insulin in both euglycemic and hyperglycemic states consistent with our earlier work and that of others (14, 25, 31). In the presence of euglycemia (group IV), there was an absolute decrease in plasma insulin in the hypoxic period compared with both hyperoxic and baseline segments. With hyperglycemia (group V), the elevated plasma insulin in segment 2 decreased markedly in segment 3 with hypoxia but only to a value that was not different from the euglycemic baseline value. The character of this inhibitory mechanism is demonstrated more clearly by the I/G during the final (hypoxic) segment, which was virtually the same with euglycemia and hyperglycemia (Fig. 4). The degree of sympathoadrenal stimulation in the two groups was equivalent as indicated by the close similarity of values for NE and Epi secretion and plasma NE. This indicates that within the observed physiological ranges, the inhibitory effect of sympathoadrenal activation is closely dependent on existing glucose-mediated insulin secretion.
It has been stated that in the adult2-adrenergic action
"inhibits insulin secretion independent of the blood glucose
concentration" (36) and furthermore that Epi and NE
completely inhibit the (insulin) secretory response to all
physiological stimuli (4). By contrast, even though this
2-inhibitory mechanism is demonstrably potent in the
fetal sheep, it does appear to be dependent on the extant glucose
concentration and does not result in an absolute cutoff of insulin
secretion as in the adult. It is possible that the degree of
hypoxia-induced sympathoadrenal activation in our in vivo fetal
preparation was less than that attained in studies in adult animals or
in vitro with adult -cells (27). It is also possible
that the inhibitory mechanism itself may be immature in the fetus. It
has been shown that 2-adrenoceptors, rather than being a
single entity, are composed of several different proteins encoded by
different genes (28). Moreover, the suggestion has been
made that different 2-receptor types may be responsible for activating different ones of the diverse intracellular events known
to result from 2-adrenergic activation in -cells. In
keeping with the concept of fetal immaturity, only a single
2-adrenoceptor type is present in islets cells of
neonatal rats (37). It may then be true that the complete
2-adrenergic inhibitory profile is not fully developed
in the sheep fetus.
- and -Adrenergic factors in control of insulin secretion.
In group VI (Table 7), idazoxan not only blocked the
insulin-inhibiting effect of sympathoadrenal activation during hypoxia but was associated with enhanced insulin secretion. This confirms that
the inhibition of insulin secretion by hypoxic stress in the sheep
fetus is based on a specific 2-adrenergic mechanism. Because experiments referred to above (groups II and
III) clearly demonstrated the absence of any agonist effect
of idazoxan on insulin, we assumed that the enhancement of insulin
secretion was due to the then unopposed action of a stimulatory
mechanism. This mechanism must of necessity have been activated by the
hypoxic stress because it was not present in the well-oxygenated fetus. This could conceivably have been parasympathetic (vagal) or
-adrenergic in nature (36). The former seems unlikely
because vagal activity in adult animals is generally believed to act
via the central nervous system to enhance insulin secretion in
anticipation of food intake, i.e., the preabsorptive insulin response.
As -adrenergic stimulation of insulin secretion has been shown (in
adult animals) to be manifest when 2-adrenergic
inhibition is blocked (27), this seemed to be the probable
explanation for the observed insulin stimulatory phenomenon. This
concept was further supported by the earlier demonstration of an active
-adrenergic stimulatory mechanism for insulin secretion in the sheep
fetus (35). This hypothesis was tested in experiments
(group VII) in which idazoxan was given in combination with
the -adrenergic blocker propranolol in hyperglycemic, hypoxic
fetuses, and in which stress-initiated sympathoadrenal activation was
not different from that in group VI. A small increase in
insulin was seen in segment 3, but insulin levels and I/G
were much lower than in experiments with idazoxan alone. The hypothesis
seems to be supported by these results, although we cannot explain the
persistence of this small insulin increase. Absence of chronotropic
responses to isoproterenol indicated that the -block was complete.
2- and -blockade.
Of significance, increases in insulin values and I/G in the low-insulin
subgroup were not different from the group VII
experiments with -blockade. This suggests that -adrenergic
stimulation of insulin secretion was absent or greatly diminished in
the low-insulin subgroup.
This phenomenon could not be explained by any consistent pattern in
fetal sex (data not shown), gestational age (data not shown), blood
oxygen levels, pH, lactate, and plasma NE, and NE and Epi secretion,
characteristic of either the low-insulin or the high-insulin subgroup.
We did observe that none of the eight fetuses in the high-insulin
subgroup had had a prior experimental hypoxic bout, whereas six of the
seven fetuses in the low-insulin subgroup had experienced one or more
2-h hypoxic bouts as part of an experiment performed 3-8
days before. It is even possible that the single fetus in the
low-insulin subgroup without a defined prior hypoxic episode may have
experienced a period of hypoxia at some time during or after surgical
preparation. Indeed, there is a wide range of insulin responses in the
high-insulin subgroup, which may reflect variations in prior incidental
hypoxic exposure. What is clear is that no fetus having
experienced a known hypoxic stress demonstrated a high-insulin response.
Even though our experiments were not designed to test for such
"residual effects" of fetal stress, our data do suggest that prior
hypoxic stress may have altered (suppressed or partially suppressed)
the -adrenergic stimulatory mechanism for insulin secretion. As
noted earlier, we found no effect of prior hypoxic exposure on the
insulin response to glucose nor on the glucose-insulin results in any
other experiment. This suggests that the apparent phenomenon of
residual suppression of -adrenergic stimulation of insulin secretion
is a specific effect acting at a point outside the primary glucose
stimulus-secretion pathway.
In summary, we have defined aspects of control of insulin secretion in
the late-gestation sheep fetus. The fetus mounts a brisk insulin
response to exogenous glucose. However, the resultant change in the I/G
is minimal, indicating a tight coupling between insulin and glucose
that is maintained in the face of continuously elevated glucose with
little if any change for a period of at least 2 h. In
well-oxygenated fetuses, 2-adrenergic blockade effects
no change in fetal plasma insulin concentrations, indicating the
absence of a resting sympathetic inhibitory tone for insulin secretion.
Hypoxic stress acting through an 2-adrenergic mechanism induces a marked but incomplete inhibition of insulin secretion, which
is quantitatively related to extant glucose concentrations. When
2-adrenergic inhibitory activity is blocked, a potent
-adrenergic stimulatory mechanism for insulin secretion becomes
manifest. Based on an analysis of the data at the completion of the
study, there is a strong suggestion that severe hypoxic stress in the fetus may, at least in the short term, have a residual effect in
suppressing this -adrenergic stimulatory mechanism.
Perspectives
Clinical studies have indicated that intrauterine events may have residual effects on glucose-insulin metabolism perinatally in the newborn (12) or many years later in the adult (18). Experimentally, glucose intolerance has been reported in the mature offspring of streptozotocin-diabetic rats (1). These clinical and experimental phenomena have been associated with small neonatal size. Nutritional deprivation during pregnancy has been suggested as the principal overall causative factor in these residual manifestations of prenatal events (18). However, a recent study in streptozotocin-diabetic rats found fetal growth restriction to be dependent on attenuated placental circulation, which suggests fetal hypoxia, as well as limited substrate transport, to be of significance in this outcome (7). We previously postulated that low-weight-for-date births, at times occurring with severe maternal diabetes, may be effected at least in part by hypoxic stress, sympathetic activation, and suppression of fetal insulin secretion with a decrease in the net growth promoting effects of that hormone (25). In support of this concept, an inverse correlation between birth weight and plasma NE was reported in neonates of diabetic mothers (38). More recently, an association was found between fetal growth retardation and maternal snoring with presumed sleep apnea (15). Here, the insult to the fetus would more likely be intermittent hypoxia rather than nutritional deprivation. The present work supports the concept of suppression of fetal insulin secretion by intrauterine events such as hypoxic stress and suggests the possibility that moderately severe hypoxia may have a residual effect in suppressing the-adrenergic stimulatory mechanism
for insulin secretion. Such an alteration, if persistent, could
exaggerate the insulin-suppressive effects of stress-initiated
sympathetic activation throughout life. The recent report of an
increased incidence of diabetes in hypertensive patients treated with
-blockers lends support to this latter concept (17).
These findings suggest the importance of further studies of fetal
hypoxic stress and consequent sympathetic activation on metabolic
events in the fetus and as causative factors in the residual effects of
pathological events during intrauterine development.
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ACKNOWLEDGEMENTS |
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We are grateful to Joan Deutsch, Nancy L. Gelardi, and Dr. Mohamed H. Traore for excellent technical assistance.
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FOOTNOTES |
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This work was supported by the National Institute of Diabetes and Digestive and Kidney Diseases Grant DK-39178.
Address for reprint requests and other correspondence: B. T. Jackson, Brown Univ. School of Medicine, Box G-B4, Providence, RI 02912 (E-mail: jackson.benjamin{at}providence.va.gov).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 29 September 1999; accepted in final form 1 August 2000.
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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