86
Neuroscience Letters, 129 (1991) 86-90 © 1991 ElsevierScientific Publishers Ireland Ltd. 0304-3940/91/$ 03.50 ADONIS 030439409100406U
NSL 07922
Cardiorespiratory responses to chemical stimulation of the caudalmost ventrolateral medulla in the cat G . A . I w a m o t o 1'2, R . D . B r t v a 1 a n d T . G . W a l d r o p 2 1Department of Veterinary Biosciences and 2Department of Physiology and Biophysics, University of Illinois, Urbana-Champaign, Urbana, IL 61801 (U.S.A.)
(Received4 February 1991; Revised version received 18 March 1991;Accepted 2 May 1991) Key words: Bloodpressure; Ventilation; Brainstem; Ventrolateral medulla; Cat
Chemical stimulation of caudal ventrolateral medulla evoked both pressor and depressor responses. The pressor sites were generally located caudal to depressor sites. Effects on heart rate were variable. Significant increases in minute ventilation were also observed, which were primarily due to changes in respiratory frequency.
The ventrolateral medulla (VLM) of the cat brain has long been identified as a pressor region. Early studies utilizing electrical stimulation (STIM) showed that an extensive ventrolateral area extending for the length of the medulla was involved in raising blood pressure [1, 19]. Recent data from m a n y species have identified cells in the rostral V L M (RVLM) which project to the spinal cord intermediolateral cell column and mediate pressor responses [3, 5, 6, 18, 20]. Recent data for the caudal ven,trolateral medulla (CVLM) are less uniform. Activation of the C V L M of the rat by electrical or chemical STIM evokes depressor responses [2, 14, 22]. However, a chemical S T I M study by G o r d o n and McCann [10] has revealed a pressor area in the rat which is caudal to the C V L M depressor area. While the caudalmost C'VLM of the cat was identified with pressor responses in older studies in using electrical S T I M [1], these data could be interpreted as stimulating fibers en passage to the R V L M . We therefore sought to determine if the cat would also show pressor responses to chemical S T I M of the caudalmost V L M as this is held to activate cell bodies while sparing axons en passage [9]. Some of these results have appeared in preliminary form [13]. Adult cats (2.3-4.0 kg, n = 22) were used in this study. Anesthesia was induced with 1-3% halothane in a mixture of 1:3 oxygen and nitrous oxide. The trachea was intubated. Arterial pressure (AP) was measured through a c o m m o n carotid artery cannula. A venous cannula was Correspondence: G.A. lwamoto, Department of Veterinary Biosciences, Collegeof Veterinary Medicine, Universityof Illinois, Urbana, IL 61801, U.S.A.
placed in the external jugular vein. The cats were then either given ~-chloralose (60 mg/kg, i.v., n = 3) or decerebrated at the midcollicular level (n = 19). Gaseous anesthetic was then discontinued. Ventilation measurements were made with a Fleisch pneumotachograph and a Gould Integrator. All cats were routinely monitored for end tidal CO2 which was maintained within a range of 3.5-4.5%. In most experiments we utilized a Radiometer ABL-3 blood analysis system to monitor CO2, 02, p H and bicarbonate. Body temperature was maintained within a range of 36.5-38°C. The caudal brainstem was then exposed for stereotaxic placement of glass micropipettes. The micropipettes ( 2 0 - 3 0 / t m tip diameter) were filled with L-glutamate ( G L U T ) in Ringer's solution (1 M, 500 or 100 mM), ?-aminobutyric acid (GABA) in Ringer's solution (500 mM), or a control solution, separated from either 1% Fast Green or 1% Pontamine Sky Blue 6BX dye by a droplet of mineral oil. Twelve animals were studied using 1 M G L U T , 3 with 500 m M and 7 with 100 mM. The 500 m M and 100 m M G L U T solutions were administered at p H 7.2-7.3. The pipette tips were placed on the surface o f the medulla at 1.5 m m rostral to 2.0 m m caudal to the level of the obex, 3.8-4.3 m m lateral to the midline. The pipettes were then advanced in steps of 0.5 or 1.0 m m from an initial depth of 3.0-4.0 m m from the dorsal surface. The chosen solution was then ejected using a pneumatic injection system (Picospritzer II, General Valve or PV 830, WPI) in total volumes of 1975 nl. Typically the injections were in 6-10 steps per STIM over a period of 5-7 s. Autoradiographic data suggesting that the extent of spread of 25 nl of tritiated
87
.A, Arterial Blood Pressure mmHg
Heart Rate beats/min
B 150 _rE. .
. . . . . . . .
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
lilt
50 L- . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200
~
CE
5ST
100
30 Tidal Volume ml 0 Breaths/min
15
I
30 seconds
Fig. 1. Exampleof cardiorespiratoryresponses obtained from the injectionof 20 nl of a 100 mM glutamate,solution utilizing a micropipettedriven by a pneumatic injection system. Note that blood pressure and respiratory frequencyare raised. Injection was made into the site marked by a dot just above lateral reticular nucleus (LRN) on the histology figure. 5ST, spinal tract of the trigeminal n.; 5SP, spinal nucleus of the trigeminal n.; LRN, lateral reticular nucleus; 12N, hypoglossal n.; CE, central canal; CI, nucleus centralis inferior; P, pyramidal tract; IOM, medial accessory inferior olive.
1 M G L U T to be 250 p m [7] from the site of injection (INJ) prompted us to largely limit our INJ to 20 nl. Selected STIM sites were marked after removing the pipette, ejecting the remaining chemical, replacing the pipette at the desired location and ejecting 100 nl dye. G L U T INJ were also made in two paralyzed cats (succinylcholine). The brain was finally removed and either rapidly frozen (Fast Green dye; cryostat sectioning) or placed in 4% neutral formalin (Pontamine Sky blue dye; sliding microtome with cryo stage). The locus o f the dye marks were evaluated by microscopic examination (Cresyl Violet staining). Results are expressed as mean-t-S.E.M. Statistical analysis utilized Student's t-test with P < 0.05 taken as significant. All summary data included in the text represent significant changes unless otherwise specified. Chemical STIM o f the caudal VLM led to significant changes in AP, respiratory rate and minute ventilation. Either pressor or depressor effects occurred with variable effects on heart rate (HR). Respiratory (RESP) changes occurred in a characteristic manner: Initially the tidal volume was decreased for a few RESP cycles accompanied by an increase in RESPrate, this was sometimes followed by increases in tidal volume. An example o f these data are shown in Fig. 1. The effects of STIM on mean
AP and H R are shown in summarizing diagrams. Fig. 2A shows CVLM sites investigated in 12 animals using 1 M G L U T INJ and the types of responses evoked at each site. Pressor responses averaged 17.8+3 m m H g (n = 28 sites). Depressor responses averaged -23.2 + 3.3 mmHg (n = 15 sites). Significant increases in minute ventilation occurred (839.9+42 to 1276.7+95 ml/min, n = 53 sites). This increase was caused by an increase in RESP frequency (25.2+1 to 2 8 . 7 + 6 breaths/rain). While increases in tidal volume were sometimes observed, these were statistically insignificant. The quantitative RESP results for each group are based on all sites for which measurements were made, irrespective of blood pressure response. Fig. 2B shows CVLM sites investigated using 100 m M G L U T INJ in 7 animals. Pressor responses averaged 12.3 + 1 mmHg (n = 29 sites). Depressor responses averaged - 1 6 . 7 _ 3 m m H g (n = 19 sites). Significant increases in minute ventilation were also observed with this stimulus (777.1-t-4 to 866.6+5 ml/min, n = 4 4 sites). Again, the significant increase was in the RESP frequency (25.2 + 1 to 28.7 + 2 breaths/min) but not tidal volume. Note that in all cases (Fig. 2), pressor responses were obtained at locations just dorsal to the L R N extending dorsally to an area just ventral to the trigeminal nucleus. While some pressor responses were evoked from the bor-
88
A
C +1.0 m m
Obex
~
5ST
-0.75 m m
~
LRN
-1.5 m m
Pressor Responses: • = 1" BP, J. HR: >10 mmHg • = $ BP, $ HR:
Neuroscience Letters, 129 (1991) 86-90 © 1991 ElsevierScientific Publishers Ireland Ltd. 0304-3940/91/$ 03.50 ADONIS 030439409100406U
NSL 07922
Cardiorespiratory responses to chemical stimulation of the caudalmost ventrolateral medulla in the cat G . A . I w a m o t o 1'2, R . D . B r t v a 1 a n d T . G . W a l d r o p 2 1Department of Veterinary Biosciences and 2Department of Physiology and Biophysics, University of Illinois, Urbana-Champaign, Urbana, IL 61801 (U.S.A.)
(Received4 February 1991; Revised version received 18 March 1991;Accepted 2 May 1991) Key words: Bloodpressure; Ventilation; Brainstem; Ventrolateral medulla; Cat
Chemical stimulation of caudal ventrolateral medulla evoked both pressor and depressor responses. The pressor sites were generally located caudal to depressor sites. Effects on heart rate were variable. Significant increases in minute ventilation were also observed, which were primarily due to changes in respiratory frequency.
The ventrolateral medulla (VLM) of the cat brain has long been identified as a pressor region. Early studies utilizing electrical stimulation (STIM) showed that an extensive ventrolateral area extending for the length of the medulla was involved in raising blood pressure [1, 19]. Recent data from m a n y species have identified cells in the rostral V L M (RVLM) which project to the spinal cord intermediolateral cell column and mediate pressor responses [3, 5, 6, 18, 20]. Recent data for the caudal ven,trolateral medulla (CVLM) are less uniform. Activation of the C V L M of the rat by electrical or chemical STIM evokes depressor responses [2, 14, 22]. However, a chemical S T I M study by G o r d o n and McCann [10] has revealed a pressor area in the rat which is caudal to the C V L M depressor area. While the caudalmost C'VLM of the cat was identified with pressor responses in older studies in using electrical S T I M [1], these data could be interpreted as stimulating fibers en passage to the R V L M . We therefore sought to determine if the cat would also show pressor responses to chemical S T I M of the caudalmost V L M as this is held to activate cell bodies while sparing axons en passage [9]. Some of these results have appeared in preliminary form [13]. Adult cats (2.3-4.0 kg, n = 22) were used in this study. Anesthesia was induced with 1-3% halothane in a mixture of 1:3 oxygen and nitrous oxide. The trachea was intubated. Arterial pressure (AP) was measured through a c o m m o n carotid artery cannula. A venous cannula was Correspondence: G.A. lwamoto, Department of Veterinary Biosciences, Collegeof Veterinary Medicine, Universityof Illinois, Urbana, IL 61801, U.S.A.
placed in the external jugular vein. The cats were then either given ~-chloralose (60 mg/kg, i.v., n = 3) or decerebrated at the midcollicular level (n = 19). Gaseous anesthetic was then discontinued. Ventilation measurements were made with a Fleisch pneumotachograph and a Gould Integrator. All cats were routinely monitored for end tidal CO2 which was maintained within a range of 3.5-4.5%. In most experiments we utilized a Radiometer ABL-3 blood analysis system to monitor CO2, 02, p H and bicarbonate. Body temperature was maintained within a range of 36.5-38°C. The caudal brainstem was then exposed for stereotaxic placement of glass micropipettes. The micropipettes ( 2 0 - 3 0 / t m tip diameter) were filled with L-glutamate ( G L U T ) in Ringer's solution (1 M, 500 or 100 mM), ?-aminobutyric acid (GABA) in Ringer's solution (500 mM), or a control solution, separated from either 1% Fast Green or 1% Pontamine Sky Blue 6BX dye by a droplet of mineral oil. Twelve animals were studied using 1 M G L U T , 3 with 500 m M and 7 with 100 mM. The 500 m M and 100 m M G L U T solutions were administered at p H 7.2-7.3. The pipette tips were placed on the surface o f the medulla at 1.5 m m rostral to 2.0 m m caudal to the level of the obex, 3.8-4.3 m m lateral to the midline. The pipettes were then advanced in steps of 0.5 or 1.0 m m from an initial depth of 3.0-4.0 m m from the dorsal surface. The chosen solution was then ejected using a pneumatic injection system (Picospritzer II, General Valve or PV 830, WPI) in total volumes of 1975 nl. Typically the injections were in 6-10 steps per STIM over a period of 5-7 s. Autoradiographic data suggesting that the extent of spread of 25 nl of tritiated
87
.A, Arterial Blood Pressure mmHg
Heart Rate beats/min
B 150 _rE. .
. . . . . . . .
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
lilt
50 L- . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200
~
CE
5ST
100
30 Tidal Volume ml 0 Breaths/min
15
I
30 seconds
Fig. 1. Exampleof cardiorespiratoryresponses obtained from the injectionof 20 nl of a 100 mM glutamate,solution utilizing a micropipettedriven by a pneumatic injection system. Note that blood pressure and respiratory frequencyare raised. Injection was made into the site marked by a dot just above lateral reticular nucleus (LRN) on the histology figure. 5ST, spinal tract of the trigeminal n.; 5SP, spinal nucleus of the trigeminal n.; LRN, lateral reticular nucleus; 12N, hypoglossal n.; CE, central canal; CI, nucleus centralis inferior; P, pyramidal tract; IOM, medial accessory inferior olive.
1 M G L U T to be 250 p m [7] from the site of injection (INJ) prompted us to largely limit our INJ to 20 nl. Selected STIM sites were marked after removing the pipette, ejecting the remaining chemical, replacing the pipette at the desired location and ejecting 100 nl dye. G L U T INJ were also made in two paralyzed cats (succinylcholine). The brain was finally removed and either rapidly frozen (Fast Green dye; cryostat sectioning) or placed in 4% neutral formalin (Pontamine Sky blue dye; sliding microtome with cryo stage). The locus o f the dye marks were evaluated by microscopic examination (Cresyl Violet staining). Results are expressed as mean-t-S.E.M. Statistical analysis utilized Student's t-test with P < 0.05 taken as significant. All summary data included in the text represent significant changes unless otherwise specified. Chemical STIM o f the caudal VLM led to significant changes in AP, respiratory rate and minute ventilation. Either pressor or depressor effects occurred with variable effects on heart rate (HR). Respiratory (RESP) changes occurred in a characteristic manner: Initially the tidal volume was decreased for a few RESP cycles accompanied by an increase in RESPrate, this was sometimes followed by increases in tidal volume. An example o f these data are shown in Fig. 1. The effects of STIM on mean
AP and H R are shown in summarizing diagrams. Fig. 2A shows CVLM sites investigated in 12 animals using 1 M G L U T INJ and the types of responses evoked at each site. Pressor responses averaged 17.8+3 m m H g (n = 28 sites). Depressor responses averaged -23.2 + 3.3 mmHg (n = 15 sites). Significant increases in minute ventilation occurred (839.9+42 to 1276.7+95 ml/min, n = 53 sites). This increase was caused by an increase in RESP frequency (25.2+1 to 2 8 . 7 + 6 breaths/rain). While increases in tidal volume were sometimes observed, these were statistically insignificant. The quantitative RESP results for each group are based on all sites for which measurements were made, irrespective of blood pressure response. Fig. 2B shows CVLM sites investigated using 100 m M G L U T INJ in 7 animals. Pressor responses averaged 12.3 + 1 mmHg (n = 29 sites). Depressor responses averaged - 1 6 . 7 _ 3 m m H g (n = 19 sites). Significant increases in minute ventilation were also observed with this stimulus (777.1-t-4 to 866.6+5 ml/min, n = 4 4 sites). Again, the significant increase was in the RESP frequency (25.2 + 1 to 28.7 + 2 breaths/min) but not tidal volume. Note that in all cases (Fig. 2), pressor responses were obtained at locations just dorsal to the L R N extending dorsally to an area just ventral to the trigeminal nucleus. While some pressor responses were evoked from the bor-
88
A
C +1.0 m m
Obex
~
5ST
-0.75 m m
~
LRN
-1.5 m m
Pressor Responses: • = 1" BP, J. HR: >10 mmHg • = $ BP, $ HR:
