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Neurons of the rostral fastigial nucleus are responsive to cardiovascular and respiratory challenges
Journal of the Autonomic Ner~,ous 5~vstem, 27 (1989) 101 112 Elsevier
llll
JANS 00939
N e u r o n s of the rostral fastigial nucleus are responsive to cardiovascular and respiratory challenges * L.O. Lutherer, J.L. W i l l i a m s ** and S.J. Everse Departments of Physiology and Internal Medicine, Texas Tech Uni~ersitv Health Sciences Center, Lubbock, TX 79430 [ U.S.A.)
(Received 29 March 1988) (Revised version received 5 April 1989) (Accepted 21 April 1989)
Key, words: Cerebellum; Extracellular recording; Blood pressure; Control of circulation: Control of respiration Abstrac!
The rostral fastigial nucleus (rFN) of the cerebellum has been implicated in the neural control of the cardiovascular and respiratory systems. Electrical stimulation and electrolytic lesions of this region produce changes in both cardiovascular and respiratory function. It has been suggested that some of these changes may result from effects on fibers of passage rather than on cell bodies of origin within the rFN. In the present study, extracellular recordings demonstrated a high percentage of units within tEN. as well as in adjacent areas, which responded to induction of acute increases or decreases in arterial blood pressure. Furthermore, units were identified in rFN which responded to respiratory stimuli as well as to changes in blood pressure. Out of the population tested, no units responding to respiratory stimuli were found in areas adjacent to rFN. In addition, a high percentage of neurons tested for response to passive movement also showed changes in firing rate to either cardiovascular or respiratory challenges, or both. Several units were identified (mostly in rFN), whose basal firing pattern was respiratory-related. This suggests the presence of cell bodies of origin within the rFN whose function is related to cardiorespiratory activity.
Introduction
the rostral fastigial nucleus ( r F N ) p r o d u c e c h a n g e s in b o t h c a r d i o v a s c u l a r a n d r e s p i r a t o r y f u n c t i o n .
Electrical stimulation of, or specific, bilateral electrolytic lesions in, the ventromedial portion of
Acute,
specific,
bilateral
lesions
of
the
rFN
markedly impair recovery of blood pressure after induction of different types of hypotension
[13,
19,37], d e c r e a s e r e s t i n g h e a r t r a t e [47] a n d a l t e r * Preliminary findings from this study were presented at the 70th Annual Meeting of the Federation of American Societies for Experimental Biology, St. Louis, MO. U.S.A., April, 1986. ** Present address: Department of Internal Medicine, Cardiovascular Division, University of Iowa College of Medicine, Iowa city, IA 52242, U.S.A. Correspondence: L.O. Lutherer, Department of Physiology, Texas Tech University Health Sciences Center, Lubbock, TX 79430, U.S.A.
baroreceptor
s e n s i t i v i t y [13]. C h r o n i c
lesions of
the r F N have b e e n r e p o r t e d to r e d u c e the p r e s s o r response and cardioacceleration observed during e x e r c i s e [21]. A c u t e l e s i o n s a l s o p r o d u c e a m a r k e d r e s p i r a t o r y d e p r e s s i o n w h i c h p e r s i s t s in s p i t e o f c h a n g e s in a r t e r i a l b l o o d g a s e s w h i c h s h o u l d p r o v i d e a n i n c r e a s e d d r i v e [47]. I n d e e d , t h e r e s p o n s i v e n e s s to a c a r b o n d i o x i d e c h a l l e n g e is m a r k e d l y d e p r e s s e d b y t h e s e l e s i o n s [39]. In c o n t r a s t , e l e c t r i -
0165-1838/89/$03.50 ~ 1989 Elsevier Science Publishers B.V. (Biomedical Division)
V)2
cal stimulation in this region activates cardiovascular systems responsible for a pressor response and increased heart rate [1,15,18,34,40] and produces changes in many respiratory parameters and influences several central respiratory systems [9,28,38,48]. These techniques, however, do not permit determination of whether these changes are due to effects on cell bodies of origin within or fibers of passage traversing this region. Recent studies [9,14,41,45] have suggested that the pressor response elicited with electrical stimulation of this area may result from activation of fibers of passage. However, the results of those studies are contradictory. It is known that neurons in this region do respond to cutaneous, somatic and vestibular afferent stimulation [3,4,6,7,22-25,27, 30,35,36], but their possible responses to activation of cardiovascular and respiratory reflexes have never been studied. The present studies, using extracellular recordings from units within the rFN, were done to determine whether general stimulation of the cardiovascular and respiratory systems could elicit changes in firing patterns in units within this region. Such responses were obtained, suggesting that there are neurons within the rFN which indeed are involved in some aspect of cardiovascular and respiratory function.
Materials and Methods
Cats of either sex (n = 9) weighing between 2.5 and 4 kg were anesthetized initially with a halothane-oxygen mixture for introduction of balloon-tipped catheters (4F) via femoral vessels into the thoracic aorta and inferior vena cava. Anesthesia was maintained subsequently by i.v. administration of chloralose-urethane (approximately 40 and 210 m g / k g , respectively). A tracheostomy was performed and a pneumotachograph (Fleisch) attached to the tracheal cannula for monitoring of tracheal airflow. In several cats, a cervical segment of the common carotid artery was isolated bilaterally for positioning of snare occluders. The animals were placed in a Kopf stereotaxic unit and a small area overlying the
cerebellum was exposed and covered with cotton soaked in warm mineral oil. Colonic lemperature was maintained between 37.5 and 38.5°C b 5 means of infrared heat lamps. The level of anesthesia was closely monitored using the jaw reflex and blood pressure and respiratory responses to noxious stimuli as indices. Supplemental anaesthesia was administered periodically to maintain a level at which reflex responses to these stimuli were not obtained. Extracellular recordings were made using stainless steel electrodes having 2-5 m~'2 impedance at 1000 Hz, 1 0 - S A (Frederick Haer). The electrodes were advanced caudorostrally at 45 ° from the vertical using a hydraulic microdrive (Trent Wells) to the predicted coordinates of the rostal FN. The raw signal was amplified (Fintronics), displayed on an oscilloscope, stored on tape for later analysis, and passed through an amplitude analyzer f o r peak discrimination. The discriminated signal was displayed on a second oscilloscope for wave form analysis and processed with a raster stepper for rate determinations. A Beckman R411 was used to record raster output, arterial blood pressure and tracheal air flow. Activity was quantified only when it could be separated clearly from background noise and demonstrated to be from a single unit. Single units were identified by the presence of a consistent wave form of constant amplitude. At the conclusion of each experiment, small DC lesions (100 /~A, 25 s) were made for histological confirmation of recording sites. Frozen sections (50 txm) were cut and stained with Cresyl violet. Basal activity was recorded from 64 cells. In addition to determining the spontaneous firing rates, the patterns were analyzed for possible respiratory-related activity. The respiratory cycle was divided into inspiratory and expiratory phases based on the tracing of tracheal air flow. The peak raster bin was determined for 20-30 consecutive respiratory cycles. Cells were classified as respiratory-related units (RRUs) if a significant number of peak bins were contained either within the inspiratory or expiratory phase as determined by a sign test. Recordings were made for 50 of the 64 units during one or more of the following cardiovascu-
103
lar a n d / o r respiratory challenges. Decreases in arterial blood pressure within the carotid sinus were induced with bilateral occlusion of the common carotid artery (CO), decreases in systemic arterial pressure by bolus infusion of sodium nitroprusside (NP, 10 ~ g / k g ) or inflation of the balloon in the vena cava (VCB), and increases in systemic pressure by bolus infusion of phenylephrine (PE, 10 /zg/kg) or inflation of the aortic balloon (AB). Respiratory challenges included intracarotid infusion of sodium cyanide (NaCN), increased tracheal dead space (TDS) and tracheal occlusion (TO). Usually, more than one of the above challenges were imposed while recording from any cell. In some cases, responses were also monitored during passive limb movement or application of air jets to the foot pads. Responses were considered positive only if the changes in neural activity exceeded 15% and similar patterns could be obtained more than once for the same challenge. Each response was reviewed several times from the signal stored on tape to insure accurate discrimination of single cells; i.e., rule out apparent changes resulting from alterations in signal amplitude rather than actual changes in discharge rate (Fig. 1).
I
2
3
4
VCB
¢ Off
Fig. 1. Oscilloscope tracing of the response d e p i c t e d for the second test in Fig. 5A. A r r o w d e s i g n a t e d VCB indicates onset of inflation of balloon in thoracic vena cava. Panels 1 4 are s e q u e n t i a l frames from the c o n t i n u o u s display of the entire r e c o r d i n g of the response to this test. H o r i z o n t a l and vertical bars represent 500 ms and 200 aV, respectively.
Results Spontaneous firing rates of cells in and around the rFN ranged from 0-90, with an average of 28.7 _+ 2.8 Hz (Table I). Histological examination revealed that 31 of these 64 neurons were in the rostral FN, The others were in the caudal FN or in the margin of the cortex immediately adjacent to the FN. One of these was identified as a Purkinje cell based on the complex discharge pattern. Often, pockets of cells were encountered in which the spontaneous activity of closely adjacent units was similar, However, there was no consistent pattern for basal activity related to anatomical location within or around the FN. Furthermore, there was no relationship between spontaneous rate and the type of response to the various stimuli subsequently applied (Table 1). Incidental identification of 3 silent cells was made by the appearance of activity coincident with tactile stimulation of the forepaws. Both regular and irregular firing patterns were observed. Some neurons displayed phasic activity, characterized by bursts of activity separated by either regular or irregular quiescent periods. The bursts ranged from several spikes to several seconds of activity, and the quiescent periods ranged from several hundred milliseconds to as long as 180 s in some cases. Seven units (5 in rostral FN) displayed phasic activity clearly correlated with the respiratory cycle (Fig. 2A) and, therefore, were classified as RRUs. Both inspiratory and expiratory neurons were identified, but no further classification was attempted. Several bins of increased activity, usually showing a ramp, were superimposed on a basal level of background firing, which continued throughout the respiratory cycle. Twenty-nine of the 31 cells in rostral FN were recorded during cardiovascular challenges, and 17 of these responded to at least one challenge (Table II). Three cells, out of the 9 tested, responded to nitroprusside-induced hypotension, each showing a marked decrease in firing rate beginning with the recovery of blood pressure (Fig. 3A). Responses to other types of cardiovascular challenges (Figs. 3 B - D and 4) included both increases and decreases in firing rate occurring with changes in arterial pressure in either direction. Although dif-
] (~4 TABLE I
Analvsis of data for spontaneous firing raters CV, c a r d i o v a s c u l a r ; R E S P , r e s p i r a t o r y ; n, number of units recorded or units responding/units tested.
A M e a n spontaneous firing rates
Spikes / s ± S. E.M.
n
All units recorded Units responding to CV stim. Units in rFN Units responding to CV stim. Units responding to RESP stim.
27.3 31.5 28.2 30.9 20.4
63 26/50 31 17/29 9/20
± ± ± ± ±
2.8 4.3 3.8 5.3 2.8
B Distribution o f spontaneous firing rates Range
A II units
A II units in r F N
C V units in r F N
R E S P units in r F N
0-10 11-20 21-30 31 50 50-90 Totals
17 8 14 14 10 63
7 5 8 6 5 31
2 4 5 3 3 17
1 4 3 1
ferent responses to the same stimulus were observed in different cells, the responses were consistent for any one cell, and more than one unit displayed each type of response. Responsiveness of some units was dependent on either the initial blood pressure or the extent of change in pressure. The firing rate of the unit depicted in Fig. 4A was markedly increased when pressure began to fall from an elevated level (release of AB) but did not change when pressure was decreased from normal (inflation of VCB). The opposite was true for the unit depicted in Fig. 4B. When units were tested at several different stimulus intensities (quantified by the change in blood pressure), it was apparent
9
that there was a threshold for obtaining a response, and the amount of response was proportional to stimulus intensity. Recordings were made from 20 of the 31 neurons within the FN during respiratory challenges, and 9 of these displayed altered firing rates (Table II). Five also responded to cardiovascular stimuli. Representative responses to N a C N and tracheal occlusion are shown on Fig. 2. Also demonstrated in Fig. 2B is altered firing in an R R U during respiratory stimulation. There was a reduction in the number of bins showing an increased activity during inspiration, and the level of background activity was reduced. Emergence of respiratory-re-
T A B L E I1
Summary of responses o f cells in r F N to oarious stimuli * CV, cardiovascular stimulation; RESP, respiratory stimulation; MOVE, passive m o v e m e n t
N u m b e r of cells responding N u m b e r of cells not responding N u m b e r of cells tested
CV
RESP
CV + RESP
MOVE
MOVE + C V or R E S P
17 12 29
9 11 20
5 15 * * 20
9 2 11
8 *** 1 9
* Not all cells were tested for each category of stimulus or for each type of stimulus within a category. * * Ten responded to either CV or RESP but not to both. * * * Four responded to both CV and RESP, one only to CV, and 3 only to R E S P .
105
mp t
t,
,
li
r /j,,j
i
AIRFLOW EXP
[~IGHARGE ImlEO
|A C 2 s*c
mH TO
~CN
Fig. 2. Respiratory-related activity of units in the rFN. Tracheal airflow is depicted by the upper tracings, raster stepper output of discharge frequency (discharges/s) by the lower tracing. A: basal firing pattern of a respiratory-related unit (RRU} located in rFN. B: response of same unit to intracarotid injection of cyanide (arrow designated NaCN). C: emergence of respiratory-related activity in a n o n - R R U during tracheal occlusion (TO).
lated activity in a n o n - R R U during the challenge is depicted in Fig. 2C. Bins with peak activity clearly associated with inspiratory activity became apparent with the onset of tracheal occlusion, and persisted after the occlusion was removed. Respiratory-related activity became apparent during respiratory challenge or passive movement in 3 cells in the rFN, that had not been classified previously as RRUs based on spontaneous activity. Eleven units within rFN were tested with passive movement and at least one other challenge.
Of the 9 cells that responded to passive movement, 3 also responded to respiratory, one to cardiovascular, and 4 to both of these additional challenges. Fifteen of the 21 units located outside tile rostral FN were recorded during respiratory stimulation, and none responded. In contrast, 9 of 21 responded to cardiovascular stimuli. Responses presented in Fig. 5 are from 5 of these cells, including one Purkinje cell (Fig. 5C). The magnitude of response of most of these cells often was greater than that of units of the rFN. A cessation
106
IB
A DISCHARGE FREQUENCY (Hz)
5O
0 200 -
BLOOD PRESSURE (ram Hg)
CO
t t l | ~ u i u ~ l l l ~ t t J ~ J • J i J ~iaJJ JJd
TRACHEAL AIRFLOW
..r.mr
lllrrll,rT r Yrrr
rlll
1 min
NP
C
IlqlllHIPHlllllllll~!lH~qIH
CO
CO
Fig. 3. Responses of units in rFN to various challenges: (A) i.v. nitroprusside (NP) and ( B - D ) bilateral carotid occlusion (CO). Decreased pressure in the carotid sinus produced decreases (B) and increases (C,D) in activity of different units.
of firing, as opposed to a simple decrease in rate, occurred frequently. As with units in rFN, responses did not occur until after a certain intensity of stimulation had been reached and thereafter were proportionate to the intensity (Fig. 5).
Discussion
The present study is the first to demonstrate that there is a large percentage of units within the rostral fastigial nucleus which alter their firing rate in response to changes in cardiorespiratory function. Some of these units responded to both cardiovascular and respiratory stimuli. Further, units were found whose basal firing pattern was respiratory-related or became respiratory-related upon induction of changes in cardiorespiratory function.
The object of this study was to evaluate a possible involvement of neurons in this region in the function of the cardiovascular and respiratory systems. This was done by demonstrating the presence, in a relatively physiological preparation, of units responsive to general changes in cardiorespiratory function. It should be noted that the stimuli used in this study were relatively non-specific. Thus, a stimulus designated as cardiovascular may have had some component of a respiratory stimulus as well. However, any possible influence of the stimuli on a second system appeared not to change the qualitative neuronal response on most occasions, because many units responding to cardiovascular stimuli, for example, did not respond to respiratory stimuli. Stimulation of specific afferents in a paralyzed preparation would permit clearer separation of responses to the separate systems, and such an approach is now clearly
107
A DISCHARGE FREQUENCY (Hz)
15 10-
50
BLOOD PRESSURE (mmHg) m
m
AB
(-
VCB
~
~
AB
D-
E
AB
VCB
E
50-
0-
AB
VCB
AB
30 sec
Fig. 4. R e s p o n s e s of u n i t s in r F N to i n c r e a s e s o r d e c r e a s e s in s y s t e m i c p r e s s u r e i n d u c e d b y i n f l a t i o n of b a l l o o n s in the v e n a c a v a ( V C B ) or t h o r a c i c a o r t a (AB). T h e u n i t in A h a d s i m i l a r r e s p o n s e s to t w o successive i n f l a t i o n s of the AB. A c t i v i t y i n c r e a s e d as p r e s s u r e fell f r o m a n e l e v a t e d level, b u t n o r e s p o n s e o c c u r r e d to a d e c r e a s e f r o m n o r m a l p r e s s u r e (VCB). In c o n t r a s t , unit in B o n l y r e s p o n d e d to fall f r o m n o r m a l p r e s s u r e . R e s p o n s e in D w a s in the o p p o s i t e d i r e c t i o n f r o m the unit in B for the s a m e s t i m u l u s . C h a n g e s in u n i t activity o c c u r r e d with l a r g e i n c r e a s e s in b l o o d p r e s s u r e in s o m e u n i t s (C) a n d with o n l y small c h a n g e s in o t h e r s (E).
warrented based on the findings of the present study. The mean spontaneous firing rates observed in the current study conformed well with those recorded extracellularly by other investigators from cells in the fastigial nucleus (FN) of decerebrate or chloralose-anesthetized cats. In previous reports, these ranged from approximately 10-40 per second [3,4,23,30]. In all reports, there was a large range from cell to cell. Firing patterns have been described routinely as regular, irregular, bursting or silent. Jahnsen [31], using intracellular recordings in vitro in a slice preparation, reported that units in the cerebellar nuclei (including FN) fired spontaneously with a regular pattern at a mean frequency of 26 + 14 Hz. Eccles et al. [25] described the existence of small pockets of cells with similar rates and patterns. This was confirmed in the present study. However, beyond the
occurrence of these pockets, there apparently is no anatomical distribution of units with respect to spontaneous activity, Furthermore. the type of response is not predictable on the basis of the spontaneous activity. The finding of neurons with respiratory-related activity (RRUs) in the FN is novel, although there are several previous reports of RRUs in the cerebellar cortex [5,8,10]. Bertrand et al. [8] reported an incidence of 28%, although this may be an overestimate [43]. In the present study, 5 of 31 units (16%) recorded in rFN were RRUs. Respiratory-related activity became evident in additional cells during imposed alterations in respiration. Both the basal and imposed respiratory-related activity may represent projections from respiratory muscles, pulmonary afferents in the vagus a n d / o r part of the costal-phrenic reflex, however. it was beyond the scope of this study to investi-
108
A DISCHARGE FREQUENCY
ES
C
5,
(Hz) 200-
--f
BLOOD PRESSURE ~oo(ram Hg)
vc'-'8 vc'-B
vc'--B
D
vSB
vc"-B
VCB
E
A--~
A~
~
~
A'--~
Fig. 5. Responses of units outside rFN to changes in systemic arterial pressure induced by inflation of balloons in the vena cava (VCB) in A - C and the thoracic aorta (AB) in D and E. Inhibition of spontaneous firing during or after challenge occurred frequently in this population ( A - C and E). A threshold for stimulus intensity is evident for units in A and B. Activity of some units both decreased and increased for opposite changes in blood pressure (D).
gate the basis of this activity. Our previous stimulation and lesion studies strongly support the concept that efferent pathways from this region influence respiration [38,47,48]. Both afferent and efferent pathways exist which could subserve this function, but a respiratory role has not been investigated for any of them. Previous investigators have studied the response of fastigial neurons to somatosensory or vestibular stimulation involving taps to and pressure on foot pads [4,23-25], air jets to foot pads [22-25], passive [3] and voluntary [7] limb movement, electrical stimulation of muscle [36] or peripheral nerves [23-25,27,35], and tilt [27,30]. Several observations from those studies are similar to those of the present study. First, the response characteristics cannot be correlated with the spontaneous firing rate, the spontaneous firing pattern, or location within the rFN. Second, a cell may respond to a single type of stimulus or to several different stimuli. Third, it may respond to
the same stimulus applied to more than one receptor field. Fourth, for a given stimulus, different cells may show no change or an increase or a decrease in firing rate. Eccles et al: [22,23] found that somatosensory stimulation elicited not only tonic excitation or inhibition, but also phasic responses with both excitatory and inhibitory components. Inhibition was attributed to direct suppression of neurons in the FN by Purkinje cells, whereas excitation resulted from disinhibition from Purkinje cells or direct activation by collaterals of climbing or mossy fibers [24]. The present study is the first to demonstrate units in the rFN that are responsive to cardiovascular and respiratory challenges. Fifty-nine percent of the cells within the rostral FN responded to one or more cardiovascular stimuli, 49 % to respiratory stimuli, and 56 % responding to respiratory also responded to cardiovascular stimuli. These percentages of responsive units are similar to those reported in studies using tilt and muscle move-
109
m e n t [7,27,30,36]. Interestingly, the p e r c e n t a g e of units r e s p o n d i n g to tilt, a stimulus which can induce changes in b l o o d pressure in a d d i t i o n to a c t i v a t i n g v e s t i b u l a r afferents, was m a r k e d l y higher in the rostral than in the c a u d a l F N [27,30]. T h e previous c o n f i r m a t i o n of the presence of units r e s p o n d i n g to s o m a t o s e n s o r y a n d v e s t i b u l a r afferent i n f o r m a t i o n was consistent with k n o w n functions of the c e r e b e l l u m a n d the p a t h w a y s involved in these responses. It is k n o w n that there are extensive p r o j e c t i o n s from n e u r o n s in the r F N to regions in the reticular f o r m a t i o n a n d b r a i n stem which are k n o w n to be involved in the neural c o n t r o l of b o t h the c a r d i o v a s c u l a r a n d r e s p i r a t o r y systems [2,12,42]. Similarly, k n o w n afferents to n e u r o n s in the r F N originate from m a n y regions involved in the control of these two systems [11,16,17,26,32,33,44,46,49]. However, a question has always existed w h e t h e r the c a r d i o v a s c u l a r a n d r e s p i r a t o r y effects o b s e r v e d with electrical stimulation of, or electrolytic lesions in, the r F N involve cell b o d i e s of origin within or axons passing t h r o u g h this region. These techniques d o not permit this type of delineation, a n d the a n a t o m i c a l l y d e m o n s t r a t e d p a t h w a y s involving n e u r o n s in this region have not been tested to d e t e r m i n e if they c a r r y i n f o r m a t i o n related to c a r d i o v a s c u l a r and r e s p i r a t o r y function. Several studies have a t t e m p t e d to a d d r e s s the a b o v e question b y using chemical stimulation. D o r m e r [20], using very large injections of glutam a t e into the r F N , o b t a i n e d a p r e s s o r response. In m o r e recent studies using microinjections, Bradley et al. [9] o b s e r v e d no changes in b l o o d pressure in response to g l u t a m a t e , a n d C h i d a et al. [14] observed only a d e p r e s s o r response with c h e m i c a l s t i m u l a t i o n with kainic acid. U m e a d i [45] r e p o r t e d p r e l i m i n a r y results showing d e p r e s s o r responses with g l u t a m a t e a n d increases, decreases a n d no c h a n g e in pressure after kainate. These results w o u l d suggest that units involved in c a r d i o v a s c u lar function are p r e s e n t in this region, b u t these investigators [9,14,45] c o n c l u d e d that the pressor responses are elicited by activating fibers of passage arising from some other u n d e s c r i b e d site. M i u r a a n d T a k a y a m a [41] a t t r i b u t e d the p r e s s o r r e s p o n s e to electrical s t i m u l a t i o n to activation of a subfastigial fiber bundle. A l t h o u g h some p a t h w a y s
influencing the c a r d i o v a s c u l a r system i n d e e d m a y pass through or near the r F N , the p r e s e n t s t u d y clearly indicates the presence of n e u r o n s within r F N that r e s p o n d to changes in c a r d i o v a s c u l a r a n d r e s p i r a t o r y function. T h e m o d a l i t y of i n p u t a n d o u t p u t of the responsive cells is not known. The o b s e r v a t i o n that a given unit can r e s p o n d to m o r e than one type of stimulus a n d the diversity of responses from unit to unit for a given type of c h a n g e in the present study suggests that a c o m plex neural n e t w o r k m a y exist. A l t h o u g h this diversity of response also might reflect the type of challenges used, this diversity of response from unit to unit is consistent with what was r e p o r t e d by Eccles [22,23] for other types of s o m a t o s e n s o r y i n p u t to this region. C o n f i r m a t i o n of this a p p a r e n t c o m p l e x network a n d the responsiveness of individual units to m u l t i p l e types of afferent i n f o r m a tion s u p p o r t the suggestion m a d e years ago [29] that the c e r e b e l l u m might be involved in integrating the o u t p u t of the c a r d i o v a s c u l a r , r e s p i r a t o r y a n d s o m a t o m o t o r systems.
Acknowledgements This study was s u p p o r t e d by A m e r i c a n H e a r t A s s o c i a t i o n G r a n t - i n - A i d 83-1235 (funds c o n t r i b uted in p a r t by A m e r i c a n H e a r t A s s o c i a t i o n , Texas Affiliate), the T a r b o x Institute of T e x a s Tech University H e a l t h Sciences Center, a n d N I H T r a i n i n g G r a n t HL07289.
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Ill) 5 Ballintijn, C.M., Luiten, P.G.M. and Juch, P.J.W., Respiratory neuron activity in the mesencephalon, diencephalon. and cerebellum in the carp, J. ('omp. Physlol.. 133 (t979) 13l -139. 6 Batini, C., Bernard, J.F., Buisseret-Delmas, C. and Horcholle-Bossavit, G., Cerebellar output responses to afferent stimulation: energy consumption and unit activity in the cat fastigial nucleus, Exp. Brain Re~'., 52 (1983) 400-410.
7 Bava, A., Grimm, R.J. and Rushmer, D.S. Fastigial unit activity during voluntary movement in primates. Brain Res., 288 (1983) 371-374. 8 Bertrand, F., Hugelin, A. and Vibert, J.F., Quantitative study of anatomical distribution of respiratory related neurons in the pons, Exp. Brain Re.r, 16 (1973) 383-399. 9 Bradley, D.J., Pascoe, J.P., Paton, J.F.R. and Spyer, K.M., Cardiovascular and respiratory responses evoked from the posterior cerebellar cortex and fastigial nucleus in the cat, J. Physiol. (Loud.), 393 (1987) 107-121. 10 Brookhart, J.M., Moruzzi, G. and Snider, R.S., Spike discharges of single units in the cerebellar cortex, ,I. Neurophysiol., 13 (1950) 465-486. 11 Carleton, S.C. and Carpenter, M.B., Distribution of primary vestibular afferents in the brainstem and cerebellum of the monkey, Brain Res., 294 (1984) 281 298. 12 Carpenter, M.B. and Batton, R.R., Connections of the fastigial nucleus in the cat and monkey, In S.L Chan-Palay and V. Chan-Palay (Eds.), The Cerebellum. New Vistas. Exp. Brain Res., Suppl. 6 (1982) 250 291. 13 Chert, C.H. and Lutherer, L.O., Fastigial nucleus lesions impair recovery of blood pressure following rapid-onset, isovolemic hypotension, Fed. Proc., 44 (1985) 1728. 14 Chida, K.. ladecola, C., Underwood, M.D. and Reis, D.J., A novel vasodepressor response elicited from the rat cerebellar fastigial nucleus: the fastigial depressor response, Brain Res.. 370 (1986) 378--382. 15 Del Bo, A., Sved, A.F. and Reis, D.J., Fastigial stimulation releases vasopressin in amounts that elevate blood pressure, Am. J. Physiol., 244 (1983) H687-H694. 16 Dietrichs, E., Cerebellar cortical afferents from the periaqueductal gray in the cat, Neurosci. Lett., 41 (1983) 21-26. 17 Dietrichs, E., Divergent axon collaterals to cerebellum and amygdala from neurons in the parabrachial nucleus, the nucleus locus coerulus and some adjacent nuclei, Anat. Emb~ol., 172 (1985) 375-382. 18 Doba, N. and Reis, D.J. Changes in regional blood flow and cardiodynamics evoked by electrical stimulation of the fastigial nucleus in the cat and their similarity to orthostatic reflexes, J. Pt~vsiol. (Loud.), 227 (1972) 729-747. 19 Doba, N. and Reis, D.J., Role of the cerebellum and the vestibular apparatus in regulation of orthostatic reflexes in the cat, Circ. Res., 34 (1974) 9 18. 20 Dormer, K.J., Foreman, R.D. and Stone, H . L Glutamateinduced fastigial pressor response in the dog, Neuroscience, 2 (1977) 577-584. 21 Dormer, K.J., Modulation of cardiovascular response to
22
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37
dynamic exercise by fastigiat nucleus, J. ,U~I',". Phy.~io/, ",6, (1984) 1369-1377. Eccles, J.C., Sabah, N.H. and Taborikova, H., Response evoked in neurons of the fastigial nucleus by cutaneous mechanoreceptors, Brain Res., 35 (1971) 523 527. Eccles. J.C., Sabah, N.H. and Taborikova, ;1., Excitatory and inhibitory responses of neurones of the cerebellar fastigial nucleus, Exp. Brain Res., 19 (1974) 61 77. Eccles, J.C., Sabah, N.H. and Taborakova, H., The pathways responsible for excitation and inhibition of fastigial neurones, Exp. Brain Res., 19 (1974) 78-99. Eccles, J.C., Rantucci, T.. Sabah, N.H. and Taborakova, H., Somatotopic studies on cerebellar fastigial cells, Exp. Brain Res., 19 (1974) 100-118. Elisevich, K.V., Hrycyshyn, A.W. and Ftumerfelt, B.A., Axonal branching in the projections from the paramedian reticular nucleus to the cerebellar cortex, Neurosci. Left., 5l (1984) 195-200. Erway, L.C., Ghelarducci, B., Pompeiano, (.). and Stanojevic, M., Responses of cerebellar fastigiat neurons to stimulation of contralateral macular labyrinth receptors. Arch. ItaL BioL, 116 (1978)205 224. Farber, J.P., Expiratory effects of cerebellar stimulation in developing opossums, Am. J. P,~vsiol., 252 (1987) Rl158 R1164. Fulton, J.F., The interrelation of cerebrum and cerebellum in the regulation of somatic and autonomic function, Medicine, 15 (1936) 247-306. Ghelarducci, B., Pompeiano, O. and Spyer, K.M., Distribution of the neuronal responses to static tilt within the cerebellar fastigial nucleus, Arch. ltaL Biol., 112 (1974) 126- 141. Jahnsen, H., Extracellular activation and membrane conductances of neurones in the guinea-pig deep cerebellar nuclei in vitro. J. Physiol. (Loud.), 372 (1986) 149-168. Kotchabhakdi, N. and Walberg, F., Cerebellar afferents from neurons in motor nuclei of cranial nerves demonstrated by retrograde transport of horseradish peroxidase, Brain Res., 137 (1977) 158 163. Kotchabhakdi, N., Hoddevik, G.H. and Walberg, F., The reticulocerebellar projection in the cat as studied with retrograde transport of horseradish peroxidase, Anat. Embryol., 160 (1980) 341-359. Koyama, S., Ammons, W.S. and Manning, J.W., Altered renal vascular tone and plasma renin activity due to fastigial and baroreceptor activation, Am. J. Physiol., 239 (1980) H232- H237. Latham, A., Paul, D.H. and Potts, A.J., Responses of fastigial neurones to stimulation of a peripheral nerve, J. Physiol. (Loud.), 206 (1970) 15P. Licata, F., Occhipinti, G. and Santangelo, F., Responses of cerebellar fastigial neurons to single muscle activation, £vp. Brain Res., 60 (1985) 127-135. Lutherer, L.O., Lutherer, B.C., Dormer, K.J., Janssen, H~F. and Barnes, C.D., Bilateral lesions of the fastigial nucleus prevent recovery of blood pressure following hypotension
111
38
39
40
41
42
43
induced by hemorrhage or administration of endotoxin, Brain Res.. 269 (1983) 251 257. kutherer, E.O. and Williams, J.L., Stimulating fastigial nucleus pressor region elicits patterned respiratory responses, Am. J. PhysioZ, 250 (1986) R418-R426. Lutherer, L.O., Everse, S.J. and Lutherer. B.C., Respiratory response to carbon dioxide is decreased by acute lesions of the fastigial nucleus, F A S E B J . , 2 (1988) A511. Miura, M. and Reis, D.J., Cerebellum: a pressor response elicited from the fastigial nucleus and its relay pathway in brainstem, Brain Res., 13 (1969) 595-599. Miura, M. and Takayama, K., The site of origin of the so-called fastigial pressor response, Brain Res., 473 (1988l 352-358. Moolenaar, G. and Rucker, H.K., Autoradiographic study of brain stem projections from fastigial pressor areas, Brain Res., 114 (1976) 492 496. Netick, A. and Orem, J., Erroneous classification of neuronal activity by the respiratory modulation index, Neurosei. Lett., 21 (1981) 301 306.
44 Somona, R. and Walberg, F., Cerebellar afferents from the nucleus of the solitary tract, Neurosci. Lett., 11 (1979) 41 47. 45 Umeadi, G., Effects of kainate and glutamate on cat fastigial nucleus, The Physiologist 30 (1987) 193. 46 Waltzer, R. and Martin, G.F., Collateralizatinn of reticulospmal axons from the nucleus reticularis gigantocellularis to the cerebellum and diencephalon. A double-labelling study in the rat, Brain Res.. 293 (1984) 153 158. 47 Williams, J.L., Robinson, P.J. and Lutherer, L.O., Inhibitory effects of cerebellar lesions on respiration in the spontaneously breathing, anesthetized cat, Brain Res., 399 (1986) 224-231. 48 Williams, J.L., Everse, S.J. and kutherer, L.O., Stimulating fastigial nucleus alters central mechanisms regulating phrenic activity, Resp. PhysioL. in press, 1989. 49 Zheng, Z., Dietrichs, E. and Walberg, F.. Cerebellar afferent fibres from the dorsal motor vagal nucleus in the cat, Neurosci. Lett., 32 (1982) 113-118.
llll
JANS 00939
N e u r o n s of the rostral fastigial nucleus are responsive to cardiovascular and respiratory challenges * L.O. Lutherer, J.L. W i l l i a m s ** and S.J. Everse Departments of Physiology and Internal Medicine, Texas Tech Uni~ersitv Health Sciences Center, Lubbock, TX 79430 [ U.S.A.)
(Received 29 March 1988) (Revised version received 5 April 1989) (Accepted 21 April 1989)
Key, words: Cerebellum; Extracellular recording; Blood pressure; Control of circulation: Control of respiration Abstrac!
The rostral fastigial nucleus (rFN) of the cerebellum has been implicated in the neural control of the cardiovascular and respiratory systems. Electrical stimulation and electrolytic lesions of this region produce changes in both cardiovascular and respiratory function. It has been suggested that some of these changes may result from effects on fibers of passage rather than on cell bodies of origin within the rFN. In the present study, extracellular recordings demonstrated a high percentage of units within tEN. as well as in adjacent areas, which responded to induction of acute increases or decreases in arterial blood pressure. Furthermore, units were identified in rFN which responded to respiratory stimuli as well as to changes in blood pressure. Out of the population tested, no units responding to respiratory stimuli were found in areas adjacent to rFN. In addition, a high percentage of neurons tested for response to passive movement also showed changes in firing rate to either cardiovascular or respiratory challenges, or both. Several units were identified (mostly in rFN), whose basal firing pattern was respiratory-related. This suggests the presence of cell bodies of origin within the rFN whose function is related to cardiorespiratory activity.
Introduction
the rostral fastigial nucleus ( r F N ) p r o d u c e c h a n g e s in b o t h c a r d i o v a s c u l a r a n d r e s p i r a t o r y f u n c t i o n .
Electrical stimulation of, or specific, bilateral electrolytic lesions in, the ventromedial portion of
Acute,
specific,
bilateral
lesions
of
the
rFN
markedly impair recovery of blood pressure after induction of different types of hypotension
[13,
19,37], d e c r e a s e r e s t i n g h e a r t r a t e [47] a n d a l t e r * Preliminary findings from this study were presented at the 70th Annual Meeting of the Federation of American Societies for Experimental Biology, St. Louis, MO. U.S.A., April, 1986. ** Present address: Department of Internal Medicine, Cardiovascular Division, University of Iowa College of Medicine, Iowa city, IA 52242, U.S.A. Correspondence: L.O. Lutherer, Department of Physiology, Texas Tech University Health Sciences Center, Lubbock, TX 79430, U.S.A.
baroreceptor
s e n s i t i v i t y [13]. C h r o n i c
lesions of
the r F N have b e e n r e p o r t e d to r e d u c e the p r e s s o r response and cardioacceleration observed during e x e r c i s e [21]. A c u t e l e s i o n s a l s o p r o d u c e a m a r k e d r e s p i r a t o r y d e p r e s s i o n w h i c h p e r s i s t s in s p i t e o f c h a n g e s in a r t e r i a l b l o o d g a s e s w h i c h s h o u l d p r o v i d e a n i n c r e a s e d d r i v e [47]. I n d e e d , t h e r e s p o n s i v e n e s s to a c a r b o n d i o x i d e c h a l l e n g e is m a r k e d l y d e p r e s s e d b y t h e s e l e s i o n s [39]. In c o n t r a s t , e l e c t r i -
0165-1838/89/$03.50 ~ 1989 Elsevier Science Publishers B.V. (Biomedical Division)
V)2
cal stimulation in this region activates cardiovascular systems responsible for a pressor response and increased heart rate [1,15,18,34,40] and produces changes in many respiratory parameters and influences several central respiratory systems [9,28,38,48]. These techniques, however, do not permit determination of whether these changes are due to effects on cell bodies of origin within or fibers of passage traversing this region. Recent studies [9,14,41,45] have suggested that the pressor response elicited with electrical stimulation of this area may result from activation of fibers of passage. However, the results of those studies are contradictory. It is known that neurons in this region do respond to cutaneous, somatic and vestibular afferent stimulation [3,4,6,7,22-25,27, 30,35,36], but their possible responses to activation of cardiovascular and respiratory reflexes have never been studied. The present studies, using extracellular recordings from units within the rFN, were done to determine whether general stimulation of the cardiovascular and respiratory systems could elicit changes in firing patterns in units within this region. Such responses were obtained, suggesting that there are neurons within the rFN which indeed are involved in some aspect of cardiovascular and respiratory function.
Materials and Methods
Cats of either sex (n = 9) weighing between 2.5 and 4 kg were anesthetized initially with a halothane-oxygen mixture for introduction of balloon-tipped catheters (4F) via femoral vessels into the thoracic aorta and inferior vena cava. Anesthesia was maintained subsequently by i.v. administration of chloralose-urethane (approximately 40 and 210 m g / k g , respectively). A tracheostomy was performed and a pneumotachograph (Fleisch) attached to the tracheal cannula for monitoring of tracheal airflow. In several cats, a cervical segment of the common carotid artery was isolated bilaterally for positioning of snare occluders. The animals were placed in a Kopf stereotaxic unit and a small area overlying the
cerebellum was exposed and covered with cotton soaked in warm mineral oil. Colonic lemperature was maintained between 37.5 and 38.5°C b 5 means of infrared heat lamps. The level of anesthesia was closely monitored using the jaw reflex and blood pressure and respiratory responses to noxious stimuli as indices. Supplemental anaesthesia was administered periodically to maintain a level at which reflex responses to these stimuli were not obtained. Extracellular recordings were made using stainless steel electrodes having 2-5 m~'2 impedance at 1000 Hz, 1 0 - S A (Frederick Haer). The electrodes were advanced caudorostrally at 45 ° from the vertical using a hydraulic microdrive (Trent Wells) to the predicted coordinates of the rostal FN. The raw signal was amplified (Fintronics), displayed on an oscilloscope, stored on tape for later analysis, and passed through an amplitude analyzer f o r peak discrimination. The discriminated signal was displayed on a second oscilloscope for wave form analysis and processed with a raster stepper for rate determinations. A Beckman R411 was used to record raster output, arterial blood pressure and tracheal air flow. Activity was quantified only when it could be separated clearly from background noise and demonstrated to be from a single unit. Single units were identified by the presence of a consistent wave form of constant amplitude. At the conclusion of each experiment, small DC lesions (100 /~A, 25 s) were made for histological confirmation of recording sites. Frozen sections (50 txm) were cut and stained with Cresyl violet. Basal activity was recorded from 64 cells. In addition to determining the spontaneous firing rates, the patterns were analyzed for possible respiratory-related activity. The respiratory cycle was divided into inspiratory and expiratory phases based on the tracing of tracheal air flow. The peak raster bin was determined for 20-30 consecutive respiratory cycles. Cells were classified as respiratory-related units (RRUs) if a significant number of peak bins were contained either within the inspiratory or expiratory phase as determined by a sign test. Recordings were made for 50 of the 64 units during one or more of the following cardiovascu-
103
lar a n d / o r respiratory challenges. Decreases in arterial blood pressure within the carotid sinus were induced with bilateral occlusion of the common carotid artery (CO), decreases in systemic arterial pressure by bolus infusion of sodium nitroprusside (NP, 10 ~ g / k g ) or inflation of the balloon in the vena cava (VCB), and increases in systemic pressure by bolus infusion of phenylephrine (PE, 10 /zg/kg) or inflation of the aortic balloon (AB). Respiratory challenges included intracarotid infusion of sodium cyanide (NaCN), increased tracheal dead space (TDS) and tracheal occlusion (TO). Usually, more than one of the above challenges were imposed while recording from any cell. In some cases, responses were also monitored during passive limb movement or application of air jets to the foot pads. Responses were considered positive only if the changes in neural activity exceeded 15% and similar patterns could be obtained more than once for the same challenge. Each response was reviewed several times from the signal stored on tape to insure accurate discrimination of single cells; i.e., rule out apparent changes resulting from alterations in signal amplitude rather than actual changes in discharge rate (Fig. 1).
I
2
3
4
VCB
¢ Off
Fig. 1. Oscilloscope tracing of the response d e p i c t e d for the second test in Fig. 5A. A r r o w d e s i g n a t e d VCB indicates onset of inflation of balloon in thoracic vena cava. Panels 1 4 are s e q u e n t i a l frames from the c o n t i n u o u s display of the entire r e c o r d i n g of the response to this test. H o r i z o n t a l and vertical bars represent 500 ms and 200 aV, respectively.
Results Spontaneous firing rates of cells in and around the rFN ranged from 0-90, with an average of 28.7 _+ 2.8 Hz (Table I). Histological examination revealed that 31 of these 64 neurons were in the rostral FN, The others were in the caudal FN or in the margin of the cortex immediately adjacent to the FN. One of these was identified as a Purkinje cell based on the complex discharge pattern. Often, pockets of cells were encountered in which the spontaneous activity of closely adjacent units was similar, However, there was no consistent pattern for basal activity related to anatomical location within or around the FN. Furthermore, there was no relationship between spontaneous rate and the type of response to the various stimuli subsequently applied (Table 1). Incidental identification of 3 silent cells was made by the appearance of activity coincident with tactile stimulation of the forepaws. Both regular and irregular firing patterns were observed. Some neurons displayed phasic activity, characterized by bursts of activity separated by either regular or irregular quiescent periods. The bursts ranged from several spikes to several seconds of activity, and the quiescent periods ranged from several hundred milliseconds to as long as 180 s in some cases. Seven units (5 in rostral FN) displayed phasic activity clearly correlated with the respiratory cycle (Fig. 2A) and, therefore, were classified as RRUs. Both inspiratory and expiratory neurons were identified, but no further classification was attempted. Several bins of increased activity, usually showing a ramp, were superimposed on a basal level of background firing, which continued throughout the respiratory cycle. Twenty-nine of the 31 cells in rostral FN were recorded during cardiovascular challenges, and 17 of these responded to at least one challenge (Table II). Three cells, out of the 9 tested, responded to nitroprusside-induced hypotension, each showing a marked decrease in firing rate beginning with the recovery of blood pressure (Fig. 3A). Responses to other types of cardiovascular challenges (Figs. 3 B - D and 4) included both increases and decreases in firing rate occurring with changes in arterial pressure in either direction. Although dif-
] (~4 TABLE I
Analvsis of data for spontaneous firing raters CV, c a r d i o v a s c u l a r ; R E S P , r e s p i r a t o r y ; n, number of units recorded or units responding/units tested.
A M e a n spontaneous firing rates
Spikes / s ± S. E.M.
n
All units recorded Units responding to CV stim. Units in rFN Units responding to CV stim. Units responding to RESP stim.
27.3 31.5 28.2 30.9 20.4
63 26/50 31 17/29 9/20
± ± ± ± ±
2.8 4.3 3.8 5.3 2.8
B Distribution o f spontaneous firing rates Range
A II units
A II units in r F N
C V units in r F N
R E S P units in r F N
0-10 11-20 21-30 31 50 50-90 Totals
17 8 14 14 10 63
7 5 8 6 5 31
2 4 5 3 3 17
1 4 3 1
ferent responses to the same stimulus were observed in different cells, the responses were consistent for any one cell, and more than one unit displayed each type of response. Responsiveness of some units was dependent on either the initial blood pressure or the extent of change in pressure. The firing rate of the unit depicted in Fig. 4A was markedly increased when pressure began to fall from an elevated level (release of AB) but did not change when pressure was decreased from normal (inflation of VCB). The opposite was true for the unit depicted in Fig. 4B. When units were tested at several different stimulus intensities (quantified by the change in blood pressure), it was apparent
9
that there was a threshold for obtaining a response, and the amount of response was proportional to stimulus intensity. Recordings were made from 20 of the 31 neurons within the FN during respiratory challenges, and 9 of these displayed altered firing rates (Table II). Five also responded to cardiovascular stimuli. Representative responses to N a C N and tracheal occlusion are shown on Fig. 2. Also demonstrated in Fig. 2B is altered firing in an R R U during respiratory stimulation. There was a reduction in the number of bins showing an increased activity during inspiration, and the level of background activity was reduced. Emergence of respiratory-re-
T A B L E I1
Summary of responses o f cells in r F N to oarious stimuli * CV, cardiovascular stimulation; RESP, respiratory stimulation; MOVE, passive m o v e m e n t
N u m b e r of cells responding N u m b e r of cells not responding N u m b e r of cells tested
CV
RESP
CV + RESP
MOVE
MOVE + C V or R E S P
17 12 29
9 11 20
5 15 * * 20
9 2 11
8 *** 1 9
* Not all cells were tested for each category of stimulus or for each type of stimulus within a category. * * Ten responded to either CV or RESP but not to both. * * * Four responded to both CV and RESP, one only to CV, and 3 only to R E S P .
105
mp t
t,
,
li
r /j,,j
i
AIRFLOW EXP
[~IGHARGE ImlEO
|A C 2 s*c
mH TO
~CN
Fig. 2. Respiratory-related activity of units in the rFN. Tracheal airflow is depicted by the upper tracings, raster stepper output of discharge frequency (discharges/s) by the lower tracing. A: basal firing pattern of a respiratory-related unit (RRU} located in rFN. B: response of same unit to intracarotid injection of cyanide (arrow designated NaCN). C: emergence of respiratory-related activity in a n o n - R R U during tracheal occlusion (TO).
lated activity in a n o n - R R U during the challenge is depicted in Fig. 2C. Bins with peak activity clearly associated with inspiratory activity became apparent with the onset of tracheal occlusion, and persisted after the occlusion was removed. Respiratory-related activity became apparent during respiratory challenge or passive movement in 3 cells in the rFN, that had not been classified previously as RRUs based on spontaneous activity. Eleven units within rFN were tested with passive movement and at least one other challenge.
Of the 9 cells that responded to passive movement, 3 also responded to respiratory, one to cardiovascular, and 4 to both of these additional challenges. Fifteen of the 21 units located outside tile rostral FN were recorded during respiratory stimulation, and none responded. In contrast, 9 of 21 responded to cardiovascular stimuli. Responses presented in Fig. 5 are from 5 of these cells, including one Purkinje cell (Fig. 5C). The magnitude of response of most of these cells often was greater than that of units of the rFN. A cessation
106
IB
A DISCHARGE FREQUENCY (Hz)
5O
0 200 -
BLOOD PRESSURE (ram Hg)
CO
t t l | ~ u i u ~ l l l ~ t t J ~ J • J i J ~iaJJ JJd
TRACHEAL AIRFLOW
..r.mr
lllrrll,rT r Yrrr
rlll
1 min
NP
C
IlqlllHIPHlllllllll~!lH~qIH
CO
CO
Fig. 3. Responses of units in rFN to various challenges: (A) i.v. nitroprusside (NP) and ( B - D ) bilateral carotid occlusion (CO). Decreased pressure in the carotid sinus produced decreases (B) and increases (C,D) in activity of different units.
of firing, as opposed to a simple decrease in rate, occurred frequently. As with units in rFN, responses did not occur until after a certain intensity of stimulation had been reached and thereafter were proportionate to the intensity (Fig. 5).
Discussion
The present study is the first to demonstrate that there is a large percentage of units within the rostral fastigial nucleus which alter their firing rate in response to changes in cardiorespiratory function. Some of these units responded to both cardiovascular and respiratory stimuli. Further, units were found whose basal firing pattern was respiratory-related or became respiratory-related upon induction of changes in cardiorespiratory function.
The object of this study was to evaluate a possible involvement of neurons in this region in the function of the cardiovascular and respiratory systems. This was done by demonstrating the presence, in a relatively physiological preparation, of units responsive to general changes in cardiorespiratory function. It should be noted that the stimuli used in this study were relatively non-specific. Thus, a stimulus designated as cardiovascular may have had some component of a respiratory stimulus as well. However, any possible influence of the stimuli on a second system appeared not to change the qualitative neuronal response on most occasions, because many units responding to cardiovascular stimuli, for example, did not respond to respiratory stimuli. Stimulation of specific afferents in a paralyzed preparation would permit clearer separation of responses to the separate systems, and such an approach is now clearly
107
A DISCHARGE FREQUENCY (Hz)
15 10-
50
BLOOD PRESSURE (mmHg) m
m
AB
(-
VCB
~
~
AB
D-
E
AB
VCB
E
50-
0-
AB
VCB
AB
30 sec
Fig. 4. R e s p o n s e s of u n i t s in r F N to i n c r e a s e s o r d e c r e a s e s in s y s t e m i c p r e s s u r e i n d u c e d b y i n f l a t i o n of b a l l o o n s in the v e n a c a v a ( V C B ) or t h o r a c i c a o r t a (AB). T h e u n i t in A h a d s i m i l a r r e s p o n s e s to t w o successive i n f l a t i o n s of the AB. A c t i v i t y i n c r e a s e d as p r e s s u r e fell f r o m a n e l e v a t e d level, b u t n o r e s p o n s e o c c u r r e d to a d e c r e a s e f r o m n o r m a l p r e s s u r e (VCB). In c o n t r a s t , unit in B o n l y r e s p o n d e d to fall f r o m n o r m a l p r e s s u r e . R e s p o n s e in D w a s in the o p p o s i t e d i r e c t i o n f r o m the unit in B for the s a m e s t i m u l u s . C h a n g e s in u n i t activity o c c u r r e d with l a r g e i n c r e a s e s in b l o o d p r e s s u r e in s o m e u n i t s (C) a n d with o n l y small c h a n g e s in o t h e r s (E).
warrented based on the findings of the present study. The mean spontaneous firing rates observed in the current study conformed well with those recorded extracellularly by other investigators from cells in the fastigial nucleus (FN) of decerebrate or chloralose-anesthetized cats. In previous reports, these ranged from approximately 10-40 per second [3,4,23,30]. In all reports, there was a large range from cell to cell. Firing patterns have been described routinely as regular, irregular, bursting or silent. Jahnsen [31], using intracellular recordings in vitro in a slice preparation, reported that units in the cerebellar nuclei (including FN) fired spontaneously with a regular pattern at a mean frequency of 26 + 14 Hz. Eccles et al. [25] described the existence of small pockets of cells with similar rates and patterns. This was confirmed in the present study. However, beyond the
occurrence of these pockets, there apparently is no anatomical distribution of units with respect to spontaneous activity, Furthermore. the type of response is not predictable on the basis of the spontaneous activity. The finding of neurons with respiratory-related activity (RRUs) in the FN is novel, although there are several previous reports of RRUs in the cerebellar cortex [5,8,10]. Bertrand et al. [8] reported an incidence of 28%, although this may be an overestimate [43]. In the present study, 5 of 31 units (16%) recorded in rFN were RRUs. Respiratory-related activity became evident in additional cells during imposed alterations in respiration. Both the basal and imposed respiratory-related activity may represent projections from respiratory muscles, pulmonary afferents in the vagus a n d / o r part of the costal-phrenic reflex, however. it was beyond the scope of this study to investi-
108
A DISCHARGE FREQUENCY
ES
C
5,
(Hz) 200-
--f
BLOOD PRESSURE ~oo(ram Hg)
vc'-'8 vc'-B
vc'--B
D
vSB
vc"-B
VCB
E
A--~
A~
~
~
A'--~
Fig. 5. Responses of units outside rFN to changes in systemic arterial pressure induced by inflation of balloons in the vena cava (VCB) in A - C and the thoracic aorta (AB) in D and E. Inhibition of spontaneous firing during or after challenge occurred frequently in this population ( A - C and E). A threshold for stimulus intensity is evident for units in A and B. Activity of some units both decreased and increased for opposite changes in blood pressure (D).
gate the basis of this activity. Our previous stimulation and lesion studies strongly support the concept that efferent pathways from this region influence respiration [38,47,48]. Both afferent and efferent pathways exist which could subserve this function, but a respiratory role has not been investigated for any of them. Previous investigators have studied the response of fastigial neurons to somatosensory or vestibular stimulation involving taps to and pressure on foot pads [4,23-25], air jets to foot pads [22-25], passive [3] and voluntary [7] limb movement, electrical stimulation of muscle [36] or peripheral nerves [23-25,27,35], and tilt [27,30]. Several observations from those studies are similar to those of the present study. First, the response characteristics cannot be correlated with the spontaneous firing rate, the spontaneous firing pattern, or location within the rFN. Second, a cell may respond to a single type of stimulus or to several different stimuli. Third, it may respond to
the same stimulus applied to more than one receptor field. Fourth, for a given stimulus, different cells may show no change or an increase or a decrease in firing rate. Eccles et al: [22,23] found that somatosensory stimulation elicited not only tonic excitation or inhibition, but also phasic responses with both excitatory and inhibitory components. Inhibition was attributed to direct suppression of neurons in the FN by Purkinje cells, whereas excitation resulted from disinhibition from Purkinje cells or direct activation by collaterals of climbing or mossy fibers [24]. The present study is the first to demonstrate units in the rFN that are responsive to cardiovascular and respiratory challenges. Fifty-nine percent of the cells within the rostral FN responded to one or more cardiovascular stimuli, 49 % to respiratory stimuli, and 56 % responding to respiratory also responded to cardiovascular stimuli. These percentages of responsive units are similar to those reported in studies using tilt and muscle move-
109
m e n t [7,27,30,36]. Interestingly, the p e r c e n t a g e of units r e s p o n d i n g to tilt, a stimulus which can induce changes in b l o o d pressure in a d d i t i o n to a c t i v a t i n g v e s t i b u l a r afferents, was m a r k e d l y higher in the rostral than in the c a u d a l F N [27,30]. T h e previous c o n f i r m a t i o n of the presence of units r e s p o n d i n g to s o m a t o s e n s o r y a n d v e s t i b u l a r afferent i n f o r m a t i o n was consistent with k n o w n functions of the c e r e b e l l u m a n d the p a t h w a y s involved in these responses. It is k n o w n that there are extensive p r o j e c t i o n s from n e u r o n s in the r F N to regions in the reticular f o r m a t i o n a n d b r a i n stem which are k n o w n to be involved in the neural c o n t r o l of b o t h the c a r d i o v a s c u l a r a n d r e s p i r a t o r y systems [2,12,42]. Similarly, k n o w n afferents to n e u r o n s in the r F N originate from m a n y regions involved in the control of these two systems [11,16,17,26,32,33,44,46,49]. However, a question has always existed w h e t h e r the c a r d i o v a s c u l a r a n d r e s p i r a t o r y effects o b s e r v e d with electrical stimulation of, or electrolytic lesions in, the r F N involve cell b o d i e s of origin within or axons passing t h r o u g h this region. These techniques d o not permit this type of delineation, a n d the a n a t o m i c a l l y d e m o n s t r a t e d p a t h w a y s involving n e u r o n s in this region have not been tested to d e t e r m i n e if they c a r r y i n f o r m a t i o n related to c a r d i o v a s c u l a r and r e s p i r a t o r y function. Several studies have a t t e m p t e d to a d d r e s s the a b o v e question b y using chemical stimulation. D o r m e r [20], using very large injections of glutam a t e into the r F N , o b t a i n e d a p r e s s o r response. In m o r e recent studies using microinjections, Bradley et al. [9] o b s e r v e d no changes in b l o o d pressure in response to g l u t a m a t e , a n d C h i d a et al. [14] observed only a d e p r e s s o r response with c h e m i c a l s t i m u l a t i o n with kainic acid. U m e a d i [45] r e p o r t e d p r e l i m i n a r y results showing d e p r e s s o r responses with g l u t a m a t e a n d increases, decreases a n d no c h a n g e in pressure after kainate. These results w o u l d suggest that units involved in c a r d i o v a s c u lar function are p r e s e n t in this region, b u t these investigators [9,14,45] c o n c l u d e d that the pressor responses are elicited by activating fibers of passage arising from some other u n d e s c r i b e d site. M i u r a a n d T a k a y a m a [41] a t t r i b u t e d the p r e s s o r r e s p o n s e to electrical s t i m u l a t i o n to activation of a subfastigial fiber bundle. A l t h o u g h some p a t h w a y s
influencing the c a r d i o v a s c u l a r system i n d e e d m a y pass through or near the r F N , the p r e s e n t s t u d y clearly indicates the presence of n e u r o n s within r F N that r e s p o n d to changes in c a r d i o v a s c u l a r a n d r e s p i r a t o r y function. T h e m o d a l i t y of i n p u t a n d o u t p u t of the responsive cells is not known. The o b s e r v a t i o n that a given unit can r e s p o n d to m o r e than one type of stimulus a n d the diversity of responses from unit to unit for a given type of c h a n g e in the present study suggests that a c o m plex neural n e t w o r k m a y exist. A l t h o u g h this diversity of response also might reflect the type of challenges used, this diversity of response from unit to unit is consistent with what was r e p o r t e d by Eccles [22,23] for other types of s o m a t o s e n s o r y i n p u t to this region. C o n f i r m a t i o n of this a p p a r e n t c o m p l e x network a n d the responsiveness of individual units to m u l t i p l e types of afferent i n f o r m a tion s u p p o r t the suggestion m a d e years ago [29] that the c e r e b e l l u m might be involved in integrating the o u t p u t of the c a r d i o v a s c u l a r , r e s p i r a t o r y a n d s o m a t o m o t o r systems.
Acknowledgements This study was s u p p o r t e d by A m e r i c a n H e a r t A s s o c i a t i o n G r a n t - i n - A i d 83-1235 (funds c o n t r i b uted in p a r t by A m e r i c a n H e a r t A s s o c i a t i o n , Texas Affiliate), the T a r b o x Institute of T e x a s Tech University H e a l t h Sciences Center, a n d N I H T r a i n i n g G r a n t HL07289.
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