Injection of CCK into the Waist Area of the Parabrachial Nucleus Increases
Taste Reactivity to Intra-Oral Infusion of Quinine in Rats
1
King ,
1
Delmond
2
Maddox
Michael S.
Joseph A.
and Lance C.
1Biology Department, Stetson University, DeLand, FL 32723
2Daytona Beach Community College, Daytona Beach, FL 32120
1. Location of Injections Site
GOAL
A.
B.
Waist (W) Area
C.
PBN Rostral (R) to Waist Area
D.
Dorsal (D) to PBN
Medial (M) to PBN
SUMMARY of RESULTS
To determine the role of CCK in the waist area of the PBN in
taste reactivity to intra-oral infusion of tastants in conscious rats.
4V
• Injections of CCK into the waist area of the PBN increased
the number of gapes, total ingestive behaviors and tongue
protrusions to quinine, but not to NaCl or sucrose.
4V
4V
bc
bc
4V
INTRODUCTION
bc
bc
• Injections of CCK into the pons but outside of the waist area
did not alter taste reactivity to these tastants.
PBN Structure and Function
0.5 mm
CONCLUSION
2. CCK Injections into W alter Oromotor Responses to Q
A.
B.
Aversive Behaviors
4. Only Injections into W alter Oromotor Responses
A.
Ingestive Behaviors
40
*
30
*
20
10
40 min.
delay
0
80
CCK
Vehicle
70
60
*
50
40
30
20
40 min.
delay
10
Gapes
Total
Total
Gapes
TP
MM
Total
TP
MM
A. CCK in W (n=5) increased the number of gapes to intra-oral infusion of
quinine (Q) (*, p<0.05 CCK vs vehicle). This effect persisted for over 40 min.
B. CCK in W increased the total number of ingestive behaviors and tongue
protrusions to Q. This effect lasted less than 40 min.
Behavioral Analysis:
• oromotor behaviors were videotaped using an S-VHS system
• taste reactivity behaviors (mouth movements, tongue protrusions,
gapes, etc) were quantified (Grill & Norgren, ’78; Spector et al., ’88)
• behaviors were compared using ANOVAs and T-tests
Histology:
• animals were overdosed with sodium pentobarbital (80 mg/kg)
• perfused with PBS then 4% paraformaldehyde
• coronal sections through PBN cut on a freezing microtome
• sections Nissl stained with thionin to verify intra-PBN infusion site
20
40 min.
delay
Ingestive Behaviors
R
D
Number of Behaviors per minute
100
80
60
40
20
40 min.
delay
CCK
Vehicle
60
50
40
30
20
40 min.
delay
10
Total
TP
MM
Total
TP
MM
10
40 min.
delay
W
R
D
M
W
R
D
M
*
CCK
Vehicle
50
40
30
20
10
40 min.
delay
0
W
R
D
M
W
R
D
M
W
R
D
M
W
R
D
M
140
120
100
E.
80
Mouth Movements
40
60
40
40 min.
delay
20
0
0
*
20
60
70
CCK
Vehicle
160
30
Tongue Protrusions
0
CCK
Vehicle
*
0
M
*#
180
120
W
M
40
D.
Number of Behaviors per minute
B.
Ingestive Behaviors
D
CCK
Vehicle
Ingestive Behaviors
3. CCK Injections into W do not alter Oromotor Responses to N or S
A.
R
C.
80
Number of Behaviors per minute
(0.233ml/1min, with dH2O rinses), wait 30 min., repeat oral infusions
40
0
Subjects:
• 12 male Wistar rats, ~300g (Hilltop Laboratories)
Intra-PBN and Intra-oral Infusions:
• 3 days of recovery, 2 of adaptation to the behavioral arena
• inject 10mM CCK or vehicle into the PBN (400nl) on subsequent days
• intra-orally infuse 0.1M NaCl, 0.1M sucrose and 0.003M quinine
60
W
METHODS
placed vertically in cerebellum dorsal to PBN (Paxinos & Watson, ‘98)
• intra-oral cannulas were constructed of PE-100 tubing with a teflon
washer, placed anterolateral to the first maxillary molar and connected
to 19g stainless steel tubing affixed to the skull with dental acrylic
CCK
Vehicle
0
Total
REFERENCES
50
Number of Behaviors per minute
CCK
Vehicle
*
Number of Behaviors per minute
Number of Behaviors per minute
80
Number of Behaviors per minute
50
cerebral ventricles (Gibbs et al., ’73; Zhang et al., ’86; Schick et al., ’88).
• Peripheral administration reduces sucrose intake in intact (Gosnell & Hsaio,
’84) and decerebrate rats (Grill & Smith, ’88).
• Although taste responses in NST and perceived intensity of sucrose solutions
are not altered by peripheral CCK (Giza et al., ’90), ingestive responses to
intra-oral infusion of sucrose are reduced (Eckel & Ossenkopp, ‘94).
• Some CCK-immunoreactive neurons within the NST project to the PBN
(Herbert & Saper, ’90) and CCK and its receptors are present within the waist
area of the PBN (Kubota et al., ’83; Block & Hoffman, ’87; Mercer & Beart,
‘97)
• The functional roles of CCK in the PBN are not known.
Intra-PBN and Intra-oral Cannula Placement:
• sodium pentobarbital anesthesia (60 mg/kg, i.p.)
• stereotaxic device with non-traumatic ear bars (Stoelting)
• double guide cannula extending 5.0mm below pedestal (Plastics One)
Gapes
Aversive Behaviors
• A gut peptide that has satiety effects when injected peripherally as well as into
Total
TP
MM
Total
A. CCK in W (same rats as in 2) does not alter total ingestive behaviors nor tongue
protrusions (TP) or mouth movements (MM) to intra-oral infusion of NaCl (N).
B. CCK in W does not alter total ingestive behaviors (Total) nor tongue protrusions
(TP) or mouth movements (MM) to intra-oral infusion of sucrose (S).
TP
MM
Data from this pilot study suggest that CCK in the waist area
of the PBN increases oromotor responses to quinine.
B.
Number of Behaviors per minute
Cholecystokinin (CCK)
Number of Behaviors per minute
The classic taste responsive PBN area, the
‘waist’ region, includes the central medial
(CM) and ventral lateral (VL) subnuclei as
well as the neurons that span the brachium
conjunctivum (bc) between these areas
(Norgren & Pfaffmann, ’75; Fulwiler & Saper, ’84; Halsell & Travers, ’97).
CCK
Vehicle
30
#
20
10
40 min.
delay
0
W
R
D
M
W
R
D
M
A. & B.
CCK in W (n=5) increased gapes to Q
but CCK rostral (R; n=2), dorsal (D; n2) or medial (M, n=3) to W did not.
(*, p<0.05 compared to vehicle)
C., D. & E.
CCK in W increased the total ingestive
behaviors and tongue protrusions to Q
but injections into nearby areas did not.
(*, p<0.05 compared to vehicle)
(#, p<0.05 comparing W to all other
groups combined).
Block, C.H. and Hoffman, G.E., 1987, Neuropeptide and monoamine components of
the parabrachial pontine complex. Peptides 8:267-283.
Eckel, L.A. and Ossenkopp, K.-P., 1994, Cholecystokinin reduces sucrose palatability
in rats: evidence in support of a satiety effect. Am. J.Physiol. 267:R1496R1502.
Fulwiler, C.E., & Saper, C.B., 1984, Subnuclear organization of the efferent
connections of the parabrachial nucleus in the rat. Br. Res. Rev., 7, 229-259.
Gibbs, J., Young, R.C. and Smith, G.P., 1973, Cholecystokinin decreases food intake
in rats. J. Comp. Physiol. Psychol. 84:323-325.
Giza, B.K., Scott, T.R and Antonucci, R.F., 1990, Effect of cholecystokinin on taste
responsiveness in rats. Am. J. Physiol. 258:R1371-R1379.
Gosnell, B.A. and Hsiao, S., 1984, Effects of cholecystokinin on taste preference and
sensitivity in rats. Behav. Neurosci. 98:452-460.
Grill, H.J., & Norgren, R., 1978, The taste reactivity test. I. Mimetic responses to
gustatory stimuli in neurologically normal rats. Br. Res., 143, 263-279.
Grill, H.J. and Smith, G.P., 1988, Cholecystokinin decreases sucrose intake in chronic
decerebrate rats. Am. J. Physiol. 254:R853-R856.
Halsell, C.B., & Travers, S.P., 1997, Anterior and posterior oral cavity responsive
neurons are differentially distributed among parabrachial subnuclei in rat.
J. of Neurophys., 78, 920-938.
Herbert, H. and Saper, C.B., 1990, Cholecystokinin-, galanin-, and corticotropinreleasing factor-like immunoreactive projections from the nucleus of the
solitary tract to the parabrachial nucleus in the rat. J. Comp. Neurol.
293:581-598.
Kubota, Y., Inagaki, S., Shiosaka, S., Cho, H.J., Tateishi, K., Hasimura, E., Hamaoka,
T. and Tohyama, M., 1983, The distribution of cholecystokinin octapeptidelike structures in the lower brain stem of the rat: An immunohistochemical
analysis. Neuroscience 9:587-604.
Mercer, L.D. and Beart, P.M., 1997, Histochemistry in rat brain and spinal cord with
an antibody directed at the cholecystokininA receptor. Neurosci. Letts.
225:97-100.
Norgren, R., & Pfaffmann, C., 1975, The pontine taste area in the rat. Br. Res., 91,
99-117.
Paxinos, G., & Watson, C., 1998, The rat brain in stereotaxic coordinates (4th ed.).
New York, NY: Academic Press.
Schick, R.R., Stevens, C.W., Yaksh, R.L., Go, V.L.W., 1988, Chronic intraventricular
administration of cholecystokinin suppresses feeding in rats. Brain Res.
448:294-298.
Spector, A.C., Breslin, P., & Grill, H.J., 1988, Taste reactivity as a dependent
measure of the rapid formation of conditioned taste aversion: a tool for the
neural analysis of taste-visceral associations. Behav. Neurosci., 102, 942952.
Zhang, D.-M., Bula, W. and Stellar, D., 1986, Brain cholecystokinin as a satiety
peptide. Physiol. Behav. 36:1183-1186.
FUNDING
NSF RUI grant IBN 0090641 and NIH RO1 DC007854