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INTRODUCTION |
Glucose is the principal regulator of insulin secretion from
pancreatic beta cells in islets of Langerhans (1, 2); however, intra-islet communication through paracrine interactions may also exert
an important level of control over insulin secretion and ultimately
glucose homeostasis. For example, glucagon secreted from islet alpha
cells potentiates insulin secretion (3), whereas somatostatin secreted
from delta cells is a potent inhibitor of glucose-stimulated insulin
secretion (4). Although these paracrine interactions are well
established, the potential autocrine action of insulin upon insulin
secretion remains unclear.
Several lines of evidence support the possibility of an autocrine
action of insulin on beta cells. Insulin binds to the surface of beta
cells (5, 6), and functional insulin receptors and receptor substrates
identical to those found in peripheral tissues have been identified in
both clonal and primary beta cells (6-9). Glucose stimulation of beta
cell lines activates the beta cell insulin receptor in the same way as
application of exogenous insulin, suggesting that insulin secreted from
beta cells binds to the insulin receptor eliciting a physiological
response (9, 10). The complete physiological consequences of insulin
receptor activation of the beta cell have yet to be completely
elucidated, but at least one effect is initiation of protein synthesis
at both transcriptional and translational levels (10-12).
Although functional insulin receptors have been identified on beta
cells, the possible effects on insulin secretion mediated by beta cell
insulin receptors have not been firmly established. Several reports
have shown that glucose-stimulated insulin or C-peptide secretion from
islets or perfused pancreas is suppressed in the presence of exogenous
insulin, leading many to believe that insulin inhibits secretion in
beta cells (13-20). Under similar conditions however, some reports
have shown no effect of insulin on glucose-stimulated insulin secretion
(21-28). Furthermore, these data are difficult to interpret as direct
autocrine action of insulin because: 1) intact organs or islets possess
neuronal and hormonal regulatory mechanisms that could interact with
exogenous insulin, 2) maintenance of normoglycemia during the time
course of the experiments is often difficult because of the addition of
exogenous insulin, and 3) high glucose levels used to evoke stimulation
likely leads to substantial activation of beta cell insulin receptors
by secreted endogenous insulin, masking the effect of exogenous insulin.
Experiments with purified beta cells and beta cell lines have
also generated conflicting evidence for insulin
feedback. Glucose-stimulated insulin secretion from purified rat beta
cells was inhibited by 20% at exogenous insulin concentrations above 1 µM (28). In contrast, measurements of the effect of
insulin on C-peptide secretion in
TC3 cells failed to show direct
evidence of secretory regulation by insulin (9). Furthermore,
transfected
TC6-F7 cells in which the insulin receptor was
overexpressed showed enhanced basal and glucose-stimulated insulin
secretion, but fractional secretory levels (percentage of total
releasable cell insulin secreted) remained unchanged at all glucose
concentrations whereas cells expressing kinase negative (inactive)
insulin receptors showed decreased glucose-stimulated insulin secretion
(11). Recent studies have confirmed that manipulation of IRS-1 levels
of beta cell lines affect levels of insulin synthesis and secretion
(29, 30). These results suggest an autocrine pathway regulating one or
more of the following: insulin secretion, insulin synthesis, and
glucose sensing/utilization.
Given the ambiguities of previous experiments, we have attempted to
directly characterize the effect of exogenous insulin upon insulin
secretion from single beta cells using amperometry (31, 32). In this
technique, an amperometric electrode is positioned next to a single
cell so that released secretory product can be detected with high
sensitivity and temporal resolution. When secretion by vesicle fusion
occurs, a current spike is recorded that corresponds to quantitative
detection of packets of molecules released by exocytosis (31-33).
Secreted insulin can be detected directly using a carbon fiber
amperometric electrode that is modified with a mixed valent film of
cyanoruthenate and ruthenium oxide (32). Alternatively,
5-hydroxytryptamine (5-HT)1 secretion can be detected as a
marker of exocytosis in beta cells (33-36). In this approach, 5-HT is
allowed to accumulate into insulin containing secretory vesicles of
beta cells, and 5-HT secretion is detected using an unmodified carbon
fiber electrode. Unless stated otherwise, we used detection of 5-HT
because it generated a higher signal to noise ratio and did not suffer
from the possible interference of exogenous insulin on secretory
measurements. This methodology allowed the measurement of exocytosis
from single, isolated cells that were not affected by possible
paracrine interactions of neighboring cells, thus allowing direct
observation of the effect of insulin on regulation of beta cell exocytosis.
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MATERIALS AND METHODS |
Chemicals and Reagents--
Bovine insulin, Type XI collagenase,
HEPES, and tolbutamide were obtained from Sigma and used without
further purification. Monoclonal anti-insulin, polyclonal anti-insulin
receptor
and IgG were obtained from BioDesign
International (Kennebunk, ME) and were of rabbit origin. Unless
otherwise stated, all chemicals for islet and cell culture were
obtained from Life Technologies. All other chemicals were from Fisher
unless noted and were of the highest purity available.
Isolation and in Vitro Culture of Mouse Islets and Beta
Cells--
Islets were isolated from 20-30 g CD-1 mice following
ductal injection with collagenase and dispersed into single cells by shaking in dilute trypsin for 10 min at 37 °C (32, 37). Cells were
cultured at 37 °C, 5% CO2, pH 7.4, in RPMI 1640 containing: 10% fetal bovine serum, 100 units/ml penicillin, and 100 µg/ml streptomycin and used on days 2 to 4 after isolation.
Isolation and in Vitro Culture of Canine, Porcine, and Human
Islets and Beta Cells--
Pancreatic islets were isolated from
canine, porcine, or human pancreas using controlled collagenase
(Boehringer Mannheim) perfusion via the duct, automated dissociation,
and discontinuous Euro-Ficoll purification using the COBE 2991 blood
cell processor as described previously (38, 39). Islets were dispersed
into single cells the next day using a previously described procedure (32). Cells were cultured at 37 °C, 5% CO2 in modified
CMRL 1066 tissue culture media containing: 10% fetal bovine serum, 25 mM HEPES, 100 units/ml penicillin, and 100 µg/ml
streptomycin, pH 7.4.
Data Collection and Analysis--
Microelectrodes were
constructed that consisted of carbon fiber disks sealed
in glass micropipettes and were polished to 30-45° angle
immediately prior to use (32, 33). Amperometry was performed using a
battery to apply potential to a sodium saturated calomel electrode
(SSCE) as described previously (33). For measurements of 5-HT
secretion, dispersed beta cells were incubated in tissue culture media
containing 0.5 mM 5-hydroxytryptamine and 1 mM
5-hydroxytryptophan for 16 h at 37 °C, 5% CO2, pH
7.4 (33). Cells were used for secretion experiments immediately after
loading. Measurements requiring direct detection of insulin were
performed on beta cells that were not allowed to accumulate 5-HT prior
to experimentation, and the microelectrode was chemically modified with
a film of mixed valent cyanoruthenate and ruthenium oxide as described
elsewhere (32). For detection of 5-HT the potential at the working
electrode was 0.65 V, whereas for detection of insulin the potential
was 0.85 V. Data were low pass filtered at 100 Hz and collected at 500 Hz using a personal computer (Gateway 2000 P5-166) via a data acquisition board (Axon Instruments, DigiData 1200B). For direct measurement of insulin using chemically modified microelectrodes, data
were further high pass filtered following collection to remove the slow
component of the background current associated with detection of the
insulin stimulant, leaving the rapid current spikes unaffected.
Amperometric measurements were made by positioning microelectrodes ~1
µm from a cell and applying stimulant from a micropipette ~30 µm
from the cell as described elsewhere (32, 33). All experiments were
performed with cells incubated at 37 °C in pH 7.4 Kreb's Ringer
buffer (KRB) containing (in mM): 118 NaCl, 5.4 KCl, 2.4 CaCl2 (unless noted otherwise), 1.2 MgSO4, 1.2 KH2PO4, 3 D-glucose, and 25 HEPES
(24 NaHCO3 for insulin measurements). Stimulant solutions
(1 nM--1 µM insulin, 200 µM
tolbutamide, 17 mM glucose) were prepared by dissolving the
desired concentration of stimulant in KRB. 30 mM KCl
stimulant was prepared as above but by removing an equal concentration
of NaCl to maintain ionic strength. All means are reported ± 1 S.E. of the mean. Statistical differences between means were evaluated
using a two-tailed Student's t test.
Anti-insulin Receptor Antibody Experiments--
Mouse beta cells
were stimulated with 100 nM insulin, and exocytosis of 5-HT
was detected by amperometry to establish viability. Following
successful stimulation with 100 nM insulin, 10 nM anti-insulin receptor
was added to the
buffer and allowed to incubate for 5 min. The same cell was then
stimulated again with 100 nM insulin in the presence of the
antibody. Following insulin stimulations, the cell was stimulated with
30 mM K+ in the presence of antibody to confirm
cell viability.
Investigation of Autocrine Activation of Beta Cells--
Direct
autocrine activation of 5-HT loaded beta cells was investigated by
first establishing cells to be responsive to 100 nM insulin
stimulation by detecting exocytosis of 5-HT by amperometry. After
establishment of viability, cells were stimulated with 30 mM K+. Following K+ stimulation, 25 nM anti-insulin receptor
was added to the buffer and allowed to incubate for 5 min. Cells were again stimulated with 30 mM K+, and the number of exocytosis
events detected in the presence and absence of antibody was compared.
In a second series of experiments, 5-HT-loaded canine beta cells were
bathed in KRB of varying H+ and Zn2+
concentration. Buffer pH was adjusted by varying bicarbonate concentration to achieve the desired pH after bubbling with 5% CO2. Extracellular Zn2+ concentration was
adjusted by adding Zn2+ to the desired concentration. Ionic
strength was held constant for all solutions. Cells were then
stimulated with 200 µM tolbutamide dissolved in KRB of
matching pH and Zn2+ concentration, and 5-HT secretion was
detected by amperometry. The number of 5-HT spikes detected per
stimulation was then compared at the various pH and Zn2+ concentrations.
Membrane Potential Measurements--
Membrane potential
measurements were made in the whole-cell perforated patch configuration
at room temperature. Pipettes were pulled from borosilicate glass and
had resistances between 4 and 6 megohm. Pipette solutions contained (in
mM): 10 KCl, 76 K2SO4, 10 NaCl, 1 MgCl2, 10 HEPES, and 200 µg/ml amphotericin B, pH 7.35. Data were low pass filtered at 100 Hz and collected at 500 Hz using
Axopatch 200A patch-clamp amplifier and DigiData digitizer (Axon
Instruments, Foster City, CA).
Extracellular Calcium Dependence of Insulin Stimulation--
The
extracellular calcium requirement for insulin stimulation was
investigated using amperometry at 5-HT-loaded mouse beta cells. Cells
were bathed in KRB containing 0 mM Ca2+ and
stimulated with 100 nM insulin dissolved in KRB containing either 0 or 5 mM Ca2+. All data come from
paired experiments of cells stimulated with both 0 and 5 mM
Ca2+-containing stimulants.
Intracellular Calcium Measurements--
Beta cells were
incubated in 2 µM Fura-2/AM (Molecular Probes) dissolved
in KRB at 37 °C, 5% CO2 for 30 min. Dye solution was
then replaced with KRB, and coverslips with adherent cells were placed
into a coverslip dish for immediate use. The resulting fluorescence
from individual cells was collected at 1 Hz through a Fluar × 40 oil immersion objective (Zeiss), band pass filter (510 ± 10 nm),
and 20 µm pinhole aperture onto a photomultiplier tube using a SPEX
CMX cation measurement system and DM3000M data acquisition software
(Instruments SA).
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RESULTS |
Insulin-stimulated Insulin Secretion in Single Beta Cells--
In
view of the majority of prior results suggesting negative feedback of
insulin on insulin secretion, it was surprising to find that
application of bovine insulin to isolated mouse beta cells at
nonstimulatory (3 mM) glucose concentrations stimulated exocytosis (Fig. 1A). This
result was confined to beta cells as the cells that responded to
insulin also exhibited exocytosis when stimulated with 17 mM glucose or 200 µM tolbutamide, stimulants known to act at beta cells (n = 8) (Fig.
1B). Furthermore, the effect did not result from a
contaminant in the insulin-stimulatory solution as addition of
anti-insulin to the stimulant solution abolished the secretory response
elicited by insulin stimulation in all cases (n = 7)
(Fig. 1C). Finally, this effect is not an artifact of
measuring accumulated 5-HT instead of insulin, as it was possible to
observe secretion by direct measurement of insulin at single beta cells
that had not been allowed to accumulate 5-HT (n = 5)
(Fig. 1D). The observation of insulin-stimulated insulin
secretion is not unique to mouse cells as we observed similar
stimulatory effects on insulin secretion following stimulation with
exogenous insulin in human (n = 10), porcine
(n = 7), and canine beta cells (n = 21)
(Fig. 2).

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Fig. 1.
Detection of exocytosis at single mouse beta
cells using carbon fiber microelectrode. Detection of 5-HT upon
application of 100 nM insulin (A) and 200 µM tolbutamide (B) (same cell as panel
A). C, detection of 5-HT upon stimulation with 100 nM insulin with 100 nM anti-insulin.
D, detection of insulin upon stimulation with 100 nM insulin with chemically modified microelectrode. In all
cases, the bar under current traces represents application
of stimulant.
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Fig. 2.
Insulin-stimulated exocytosis in beta cells
from different species. Traces represent detection of
accumulated 5-HT with bare carbon fiber microelectrode from single,
dispersed beta cells following application of 100 nM
insulin dissolved in KRB as indicated by the bars underneath
the trace. Detection is from canine (A), porcine
(B), human (C), and mouse (D) beta
cells.
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Requirement of Beta Cell Insulin Receptor for Insulin-stimulated
Exocytosis--
To determine whether insulin-stimulated insulin
secretion was mediated by the beta cell insulin receptor, we examined
the antagonistic effect of anti-insulin receptor
on
insulin-stimulated insulin secretion (Fig.
3). For these experiments, cells were stimulated with 100 nM insulin to establish responsiveness,
then incubated with 10 nM polyclonal anti-insulin
receptor
to block the insulin receptor, and stimulated
again with 100 nM insulin. In no case was secretion
detected following application of antibody (n = 4).
Subsequent stimulation with 30 mM K+ in the
presence of anti-insulin receptor
evoked secretion in
all cases establishing cell viability following antibody treatment. In
control experiments, addition of immunoglobulins had no detectable effect on insulin-stimulated exocytosis (data not shown). We also found
that the insulin-stimulated insulin secretion was dependent upon the
concentration of applied insulin in the range of 1-100 nM
as illustrated in Fig. 3, E and F. It was found
that higher concentrations of insulin (1 µM) did not
induce a significantly different number of exocytosis events. (In
evaluating the concentrations used to elicit secretion, it is important
to realize that the concentrations reported are those present in the
pipette. During stimulation, the solution is diluted by ill-defined
amount as it is applied to the cell.)

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Fig. 3.
Receptor dependence of insulin-stimulated
exocytosis in single mouse beta cells. All data in panels
A-D are detection of accumulated 5-HT from same cell.
A and B, detection of exocytosis during
stimulation with 100 nM insulin at 5-min time intervals.
C, detection of exocytosis upon stimulation with 100 nM insulin following 5-min incubation in 10 nM
polyclonal anti-insulin receptor . D,
detection of exocytosis upon stimulation with 30 mM
K+ in the presence of 10 nM polyclonal
anti-insulin receptor . Bars underneath traces
represent application of stimulant. E, dependence of
secretory activity, measured as number of exocytosis events detected,
upon concentration of insulin in stimulant solution (n = 7 and 8 for 1 and 100 nM insulin, respectively) in mouse
beta cells. Statistical significance for * is p < 0.025. F, dependence of secretory activity, measured as
number of exocytosis events detected, upon concentration of insulin in
stimulant solution (n = 4, 14, and 12 for 1, 10, and
100 nM insulin, respectively) in canine beta cells.
Statistical significance for * is p < 0.05. Higher
concentrations of insulin (1 µM) did not evoke a
significantly different number of exocytosis events.
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Direct Autocrine Stimulation of Single Beta Cells--
Next we
investigated the possibility of direct autocrine action of insulin at
single beta cells. Cells that had been established as
insulin-responsive by detection of 5-HT secretion following insulin
stimulation were stimulated with 30 mM K+ in
the presence and absence of 25 nM anti-insulin
receptor
to prevent autocrine activation of the beta
cell insulin receptor. Stimulation resulted in 20.2 ± 4.5 spikes
per stimulation preceding addition of antibody and was reduced to
9.6 ± 2.4 spikes per stimulation (n = 13) upon
addition of antibody (p < 0.05), indicating that antibody could block released insulin from further enhancing release.
As further confirmation that insulin could contribute to direct
positive feedback, we investigated the effects of conditions that
reduce the free insulin concentration at the cell surface. We have
previously shown that increasing H+ and Zn2+ in
the extracellular medium significantly decreases the extrusion rate of
insulin from single secretory vesicles after vesicle fusion leading to
a decrease in the concentration of free insulin at the cell surface in
cells undergoing exocytosis (33, 34). Table
I summarizes the amount of 5-HT
secretion, measured as a number exocytosis events detected, evoked by
200 µM tolbutamide detected from cells incubated in
control buffers and buffers containing varying H+ and
Zn2+ concentrations. The number of spikes detected by
stimulation is significantly reduced at higher H+
(p < 0.01) and Zn2+ (p < 0.005) concentrations. Thus, under conditions where the amount of free
insulin is reduced at the cell surface after vesicle fusion, the number
of exocytosis events is reduced. These results are consistent with the
hypothesis that secreted free insulin activates further insulin
secretion.
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Table I
Effect of insulin complexation on autocrine feedback of beta cell
insulin receptor
Cells were stimulated with 200 µM tolbutamide, and 5-HT
secretion was detected by amperometry under control conditions and
conditions where the amount of free insulin at the cell surface was
reduced by slowing the dissociation and/or dissolution of the
zinc:insulin complex. The number of stimulations is given as
n.
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Glucose Dependence of Insulin-stimulated Insulin
Secretion--
The primary physiological action of insulin is to
stimulate glucose uptake and utilization, therefore we examined the
interaction between exogenous insulin stimulation and extracellular
glucose concentration by measuring the effect of insulin stimulation
upon insulin secretion of beta cells bathed in 0, 3, and 20 mM glucose. At 0 and 3 mM glucose, application
of 100 nM insulin evoked a similar number of exocytosis
events per stimulation, 10.3 ± 2.2, n = 8, and
7.2 ± 1.4, n = 8, for 0 and 3 mM
glucose, respectively (p = ns). In cells that were
bathed in 20 mM glucose, application of 100 nM
exogenous insulin further increased the frequency of detected
exocytosis events from 0.168 ± 0.117 s
1 to
0.216 ± 0.084 s
1 (n = 5). This
increase, however, was not statistically significant.
Effects of Insulin on Membrane Potential and Intracellular
[Ca2+]--
The novelty of insulin as an insulin
secretagogue prompted us to explore other effects of insulin on
stimulus-secretion pathways. Many insulin secretagogues such as
glucose, sulfonylureas, and K+ depolarize the plasma
membrane leading to opening of L-type voltage-gated Ca2+
channels and thereby allowing Ca2+ entry into the cell and
initiation of exocytosis. Insulin however, did not depolarize the
membrane in beta cells that could be depolarized by 200 µM tolbutamide (Fig.
4A). The average change in
membrane potential from base line to plateau (not including action
potential) following tolbutamide stimulation was 32.5 ± 1.58 mV
with the occurrence of action potentials although the average change
following insulin stimulation was 1.47 ± 1.52 mV
(n = 13). (Although some cells showed a small
depolarization, such as that shown in Fig. 4A, others had no
effect or slight hyperpolarization.) Insulin-stimulated secretion was
not dependent upon extracellular Ca2+ (Fig. 4B)
as the numbers of exocytosis events were not significantly different in
cells bathed in Ca2+-free KRB and stimulated with 100 nM insulin containing 5 mM or 0 mM
Ca2+ (n = 10). This result is not an
artifact of low levels of Ca2+ in the media as we have used
similar protocols to demonstrate the Ca2+ dependence of
glucose, tolbutamide, and K+ stimulation (32). Although
extracellular Ca2+ was not required to initiate secretion,
stimulation with insulin did cause increases in intracellular calcium
([Ca2+]i) (n = 5)
as seen in Fig. 4C. Also seen in Fig. 4C, the
insulin-evoked [Ca2+]i changes
were smaller than those caused by K+ but were of longer
duration.

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Fig. 4.
Effect of insulin on membrane potential and
[Ca2+]i in single
mouse beta cells. A, membrane potential measurements
following stimulation with 200 µM tolbutamide
(T) and 100 nM insulin (I) at a
single beta cell. B, detection of exocytosis during
application of 100 nM insulin to cells incubated in
Ca2+-free KRB. Stimulant contained 5 mM
CaCl2 or 0 mM CaCl2 as indicated.
Data are from the same cell. C, Fura-2 microfluorimetry
measurements of [Ca2+]i changes
following application of 30 mM K+ or 200 nM insulin (I) dissolved in KRB containing 2.4 mM Ca2+. For all traces, bar
represents application of stimulant for 30 s.
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DISCUSSION |
We have demonstrated for the first time that insulin can stimulate
insulin secretion in pancreatic beta cells (Fig. 1D). This effect is mediated by the beta cell insulin receptor as evidenced by
the antagonistic effect of anti-insulin receptor (Fig. 3). Furthermore,
the insulin concentrations necessary for this effect are in the
nanomolar range, which is reasonable because the EC50 of
the beta cell insulin receptor is ~4 nM (9). Insulin
released from a single cell is present at a sufficient level to
activate the receptor and enhance secretion as demonstrated by the
following results: 1) decreased secretion stimulated by K+
when the insulin receptor is blocked by anti-insulin; 2) decreased tolbutamide-evoked secretion when the cell surface concentration of
free insulin is reduced by increases in H+ and
Zn2+ extracellular concentration (Table I); and 3) the
relatively minor enhancement of secretion by exogenous insulin during
stimulation with 20 mM glucose, a condition in which
released endogenous insulin may be expected to activate receptors to
near maximal levels. Our observation that released insulin can activate
insulin autoreceptors is in agreement with previous results which
demonstrated that beta cell insulin receptors are activated by glucose
(9). Taken together, these results suggest that a portion of secretion
that is normally observed from single beta cells is because of positive autocrine feedback upon beta cell exocytosis acting through the beta
cell insulin receptor.
Although insulin evokes secretion, it does not evoke membrane
depolarization and subsequent Ca2+ entry to cause secretion
as evidenced by the minimal effect on membrane potential (Fig.
4A) and independence of the insulin-stimulatory effect on
extracellular Ca2+ (Fig. 4B). Insulin-stimulated
insulin secretion is not mediated by glucose or increases in glucose
utlization as the effect occurs even at 0 mM glucose.
Insulin does, however, evoke a rise in
[Ca2+]i. These results are
consistent with a mechanism of stimulation in which insulin evokes
release of intracellular calcium stores to initiate exocytosis
reminiscent of ATP binding to P2y receptors in beta cells
(40, 41).
Autoreceptor effects on hormone or neurotransmitter secretion are well
known; however, most autoreceptors mediate negative feedback on
secretion. The beta cell-insulin system appears to be a rare example of
positive feedback on secretion. The interplay of this positive feedback
effect with other regulatory mechanisms to control insulin secretion
and glucose homeostasis in vivo is likely complex. It is
reasonable to speculate that positive feedback would cause augmented
secretion during the initial stages of elevated glucose levels giving
rise to a greater bolus of insulin release; however, other mechanisms
must eventually take over to suppress release. Such a sequence could
contribute to the rapid increase observed in first phase insulin
secretion and the sustained, lower secretion during second phase. The
possibility that insulin has a local effect on secretion also raises
the possibility of novel regulatory mechanisms that might occur within
islets. For example, because Zn2+ and H+ can
control the cell-surface concentration of insulin after vesicular fusion (33), these ions could play a role in regulating insulin secretion by affecting positive feedback. Serum concentrations of
Zn2+ are in the range of 15-25 µM (42),
which would be sufficient to have a large effect on free insulin level
around the beta cell.
The existence of positive feedback may allow explanation of several
phenomena in beta cells. For example, cultured beta cells in contact
with other beta cells have been shown to secrete more insulin than
isolated cells (43, 44). This result has not been explained but could
be mediated by insulin from one cell stimulating further release in
neighboring cells. In addition, insulin secretion from islets has been
demonstrated to be oscillatory in nature and many models for
oscillation have assumed some form of positive feedback by a diffusible
factor released from beta cells (45, 46). No compound has been
satisfactorily identified that could serve this role; however, these
results indicate insulin as a possible candidate. Oscillations in
insulin release are of significant interest because loss of oscillatory
release is an early symptom of type-II diabetes (47). Finally, it has
been demonstrated that many type II diabetics have a marked reduction in first phase insulin secretion (48), which to date has remained unexplained but could be envisioned as involving lack of positive feedback from the beta cell insulin receptor. Supporting the
possibility that autocrine activation of the beta cell insulin receptor
is involved in early secretory responses is recent work in which mice
with knock-outs of the beta cell insulin receptor have impaired early
insulin secretion and concomitant glucose intolerance (49).
Perhaps most importantly, these results suggest a possible link between
impaired insulin secretion and insulin resistance, both of which can
lead to hyperglycemia and are considered hallmarks of type-II diabetes
(50). Considerable controversy surrounds the issue of which of these
deficiencies is the primary cause of diabetes. In some studies, the
earliest observed defect is dysfunctional secretion (45, 47) and in
others insulin resistance appears to be the first detectable problem
(51). The observation that insulin receptors on beta cells mediate
insulin secretion and synthesis (10, 11, 12), in addition to the well
known role in activating glucose utilization, leads to the possibility of a direct link between dysfunctional insulin secretion and insulin resistance. Such a link is supported by evidence that disruption of the
beta cell insulin receptor (49) or beta cell receptor substrates (11,
29, 30) induces defects in secretion whereas disruption in insulin
receptor substrates induces insulin resistance (52, 53).