Injection of CCK into the Waist Area of the Parabrachial Nucleus Increases Taste Reactivity to Intra-Oral Infusion of Quinine in Rats

Injection of CCK into the Waist Area of the Parabrachial Nucleus Increases Taste Reactivity to Intra-Oral Infusion of Quinine in Rats

Michael S. King1, Joseph A. Delmond1 and Lance C. Maddox2

1Biology Department, Stetson University, DeLand, FL 327232Daytona Beach Community College, Daytona Beach, FL 32120

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GOALGOALTo determine the role of CCK in the waist area of the PBN intaste reactivity to intra-oral infusion of tastants in conscious rats.

(Norgren & Pfaffmann, ’75; Fulwiler & Saper, ’84; Halsell & Travers, ’97).

INTRODUCTIONINTRODUCTION

PBN Structure and Function

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

Cholecystokinin (CCK)

• A gut peptide that has satiety effects when injected peripherally as well as into 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.

METHODSMETHODSSubjects:

• 12 male Wistar rats, ~300g (Hilltop Laboratories)

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) 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

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 (0.233ml/1min, with dH2O rinses), wait 30 min., repeat oral infusions

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

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CCK Injections into W alter Oromotor Responses to Q

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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.

CCK Injections into W do not alter Oromotor Responses to N or S

A. CCK in W (same rats as in 2) does not alter total ingestive behaviors nor tongueprotrusions (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).

Only Injections into W alter Oromotor Responses

Location of Injections SiteD.

A.

A. & B. CCK in W (n=5) increased gapes to Q but CCK rostral (R; n=2), dorsal (D; n-2) 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).

0.5 mm

4V

bcbc4V

bc4V

bc

4V

Waist (W) Area PBN Rostral (R) to Waist Area Dorsal (D) to PBN Medial (M) to PBN

FUNDINGFUNDINGFUNDINGFUNDINGNSF RUI grant IBN 0090641 and NIH RO1 DC007854

REFERENCESREFERENCESREFERENCESREFERENCES

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:R1496- R1502.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

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Herbert, H. and Saper, C.B., 1990, Cholecystokinin-, galanin-, and corticotropin-releasing 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 octapeptide- like 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

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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, 942-

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CONCLUSIONCONCLUSIONCONCLUSIONCONCLUSIONData from this pilot study suggest that CCK in the waist areaof the PBN increases oromotor responses to quinine.

SUMMARY of RESULTSSUMMARY of RESULTSSUMMARY of RESULTSSUMMARY of RESULTS• 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.

• Injections of CCK into the pons but outside of the waist area did not alter taste reactivity to these tastants.