Dorsal raphe and nociceptive stimulations evoke convergent responses on the thalamic centralis lateralis and medial prefrontal cortex neurons

Brain Research, 499 (1989) 145-152

145

Elsevier BRES 23736

Dorsal raphe and nociceptive stimulations evoke convergent responses on the thalamic centralis lateralis and medial prefrontal cortex neurons Miguel Cond6s-Lara 1, Imelda Omafia Zapata 3, Martha Le6n-Olea 2 and Marcela S~inchez-Alvarez 2 l Departamento de Neurofisiologfa, 2Laboratorio de Histologfa, Divisi6n de Investigaciones en Neurociencias, Instituto Mexicano de Psiquiatria, M~xico (Mexico) and 3Coordinaci6n de Estudios de Postgrado, lnvestigaci6n y Desarrollo Acad~mico ENEP, Zaragoza, UNAM, M~xico (Mexico)

(Accepted 13 June 1989) Key words: Nociception; Medial prefrontal cortex; Centralis lateralis nucleus; Dorsal raphe nucleus; Ascending pain modulation

pathway; Rat

There is evidence for the existence of a descending pain suppression system, but also there are data supporting the hypothesis for the modulation of pain at higher central nervous system levels. In the present study we give evidence for a possible ascending pain modulation pathway which involves the dorsal raphe (DR), the centralis lateralis nucleus (CL) of the thalamus and the medial prefrontal cortex (PFCx). Urethane-anesthetized rats were used. Simultaneous single unit recordings were done in the CL and PFCx regions under noxious and DR stimulations. Cells responding to both types of stimuli exhibit duration responses directly related to the duration of the stimuli. Thus, from our results we conclude a DR influence upon CL and PFCx structures that are involved in the coding of nociceptive information. A possible route for an ascending pain modulation path is proposed.

It is well established that the electrical stimulation of m i d b r a i n structures elicits behavioral analgesia and decreases the e v o k e d nociceptive discharges in dorsal horn cells and spinothalamic pathways 12' 23,26,27,31 H o w e v e r , there is evidence that pain m o d u l a t i o n also occurs at higher CNS levels. For e x a m p l e , the parafascicularis nuclei (Pf) and nearby thalamic regions have been shown to be responsive to noxious stimuli in the m o n k e y , cat and rat (reviewed in ref. 2), and dorsal r a p h e ( D R ) stimulation m o d u l a t e s the noxious input in Pf neurons 7' 8,32. These findings suggest the possible existence of an ascending pain modulation system by way of a supraspinal p a t h w a y 6'7'32 in addition to the described descending paths. A t the present, the role of spatially recognizing and discriminating nociceptive sensations is attributed to the ventrobasal complex and its projections

to the s o m a t o s e n s o r y cortex, whereas the unpleasant feelings associated with pain and the control o f m o t o r responses to nociceptive stimuli is attributed to the medial thalamus 2,22,39. The centralis lateralis nucleus of the thalamus (CL) seems to play a particular role in mechanisms associated with nociception, because it is the only area of the intralaminar complex which receives a dense spinothalamic and trigeminothalamic inputs TM 12,24,30 Also, CL neurons exhibit responses to noxious p e r i p h e r a l stimulation 13,14,29. Recently 14, we d e m o n s t r a t e d a cortical facilitatory control upon the noxious responses r e c o r d e d in the C L cells. The action of prefrontal cortex (PFCx) has also been shown to play a role in pain m o d u l a t i o n 6' 10.19-21 and cells in this area exhibited responses to noxious stimulation 14. T h e PFCx could m o d u l a t e responses of thalamic and m i d b r a i n neurons to

Correspondence: M. Cond6s-Lara, Departamento de Neurofisiologfa, Divisi6n de Investigaciones en Neurociencias, Instituto Mexicano de Psiquiatria, Calzada M6xico-Xochimilco no. 101, M6xico, D.F. 14370, Mexico.

0006-8993/89/$03.50 (~) 1989 Elsevier Science Publishers B.V. (Biomedical Division)

146 noxious stimulation 6"14"21. Moreover, in an accompanying paper we give electrophysiological evidence for pathways originating in DR and reaching the CL and PFCx 15. Also, the PFCx and the CL are reciprocally connected 4,5,m'36. The electrical stimulation of the thalamus such as the centromedian-parafascicular complex relieves chronic pain 9"33"3s, and it is suggested that the analgesic effects of prefrontal thalamic stimulation is due to activation of the descending pain suppression system by exciting the periaqueductal gray (PAG) and adjacent reticular formation (RF) neurons 34. The aim of the present paper is to give some evidence about the properties of the CL and PFCx cells codification of nociceptive and DR input. Twenty-six rats (Wistar) weighing 250-300 g were used to observe the effects produced by mesencephalic electric and nociceptive thermic stimulation upon CL and PFCx extracellular unit records. Subjects were anesthetized with urethane (15002000 mg/kg i.p.) and placed in a stereotaxic apparatus. The stereotaxic coordinates and the incisor bar orientation were those used by Albe-Fessard et al. 5. The head was shaved and the skull exposed. Two holes were drilled into the bone. One was used to insert a concentric bipolar stainless steel electrode (0.3 mm tip contact separation) into the mesencephaion at a 20 ° angle lateral from the midline. This procedure permitted the electrode to avoid the superior sagittal sinus. The bipolar electrode was used for electrical stimulation and was placed at coordinates of interaural 2.2 to -0.8, lateral 0.0 using the Paxinos and Watson 2s stereotaxic coordinates. The stimulation parameters were 0.5 ms biphasic square pulses (0.1 ms between them), 20 Hz and 0.1-1.5 mA. The train duration varied from 0.2 to 5 s. The other hole, made between A 4.5 and 10.5, and L 0.5 to 2.5 (ref. 5), allowed us to place the electrodes for recording unit activity. We studied cells located in the PFCx and in the CL of the thalamus. Single units were recorded using glass micropipettes filled with a 4% solution of Pontamine sky blue in 1 M KC! (impedance 8-10 Mg2). A systematic mapping of CL and PFCx cells was performed and the final recording sites were stained by iontophoretic injection of Pontamine blue (15/~A cathodal current applied during 30 min). The tip location of the stimulation electrode was possible

by using a 5-mA DC current for a 5-s period. The animals were overdosed with pentobarbital and perfused with 10% formalin and 3% potassium ferrocyanide to produce a Prussian blue spot at the site of the stimulation electrode. The brains were cut (50 ~m) and the sections were observed after Nissl staining. The electrode tracks were then reconstructed using microdrive reference points. During the entire experiment, heart rate was monitored continuously and body temperature maintained at about 38 °C by a hot water circulating pad. We tested the responses of thalamic and cortical cells to noxious heat stimulation that consisted of immersing the rat's tail in water at 50 °C for times between 10 and 50 s. Thalamic and cortical unit activity were recorded simultaneously using two separate amplification channels. Unit activities were digitalized and counted. These counts were plotted automatically by frequency: impulses per second. Also, unit activity was stored in a Personal Computer and off-line autocorrelation histograms were constructed. Noxious stimulation was delivered with at least 20 rain elapsing between tests. PFCx and CL cell responses were simultaneously studied during nociceptive and DR stimulation. The 20-min intertrial periods between noxious stimulations were intended to reduce possible accumulative effects, or receptor adaptation processes. The 20-min intertrial periods required us to record a unit's activity for periods longer than 30 min. We studied 93 units localized in the CL and 62 in PFCx, during D R or median raphe nucleus (MnR) electrical stimulation. Afterwards, the responsive cells were tested for noxious thermic stimulation (50 °C). Out of 80 CL cells that responded to D R stimulation, only 40 exhibited responses to the noxious stimulation of the tail (50 °C). Of a total of 44 PFCx responsive cells to D R stimulation, 18 of them exhibited responses to noxious stimulation. CL and PFCx cells could respond with an increase or a decrease in their firing rate to D R stimulation. Only cells exhibiting increases in their response to DR were also responsive to noxious stimulation. Cells in CL and PFCx which were responsive to DR and 50 °C stimulation, were called 'convergent' cells. These data are concentrated in Table I.

147 TABLE I

durations directly related to the duration of 50 °C or D R stimulations. Typical responses of an identified CL cell to 50 °C (Fig. 1A) and D R (Fig. 1B,C) stimulations are illustrated in Fig. 1. The convergent attribute in cell responses to D R and 50 °C stimulations was also manifested by a proportional dura-

CL and PFCx cellular responses to dorsal raphe (DR), noxious (50 °C) stimulations, and those responding to both types o f stimuli (50 °C, DR)

( I' ) firing rate increased, ( $ ) firing rate decreased. Note the lack of CL and PFCx decreased responses to 50 °C stimuli. CL

tion increase in frequencies for different durations of

PFCx

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80 47 40 93

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responses was directly related with the stimulus duration for both types of stimulus (Fig. 1A,B). D R stimulus with the same duration produced similar duration of responses (Fig. 1C). D R stimulation not only produced increases or

The spontaneous neuronal activity of CL and PFCx cells was characterized by a low firing rate

decreases in the firing rate of the PFCx cells, but there were also changes in the pattern of discharge. Fig. 2A illustrates the autocorrelation histograms of

(1-5 Hz), and exhibited increases in its activity with

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Fig. 2. PFCx cell responses to DR and 50 °C stimulations. In A, the upper part shows the autocorrelation histograms before (at the left) and after DR stimulation (at the right). In the middle part is the time-frequency histogram before and after 5 s of DR stimulation (St Raphe), this histogram used a bin time of 1 s. On the bottom, the traces of the pattern of neuronal firing: spontaneous at the left, and after DR stimuli at the right. In B, the same cell responding to DR stimulation (2 and 3 s) and to 50 °C stimulation. Note the rhythmic pattern produced by DR stimulation of about 5 cycles per s (in the upper right side of the figure). Numbers below dots and bars represent the duration of stimulation and time calibration.

a cortical cell before and after 5 s of D R stimulation. D R stimulation p r o v o k e d a slight increase in the neuronal firing rate and also p r o d u c e d a rhythmic burst discharge of about 5 Hz. Rhythmic responses were observed in 5 of a total of 44 DR-responsive PFCx cells. This type of cortical cells also exhibit a convergent response to D R and 50 °C stimulations (Fig. 2B). Thalamic and cortical cells were classified according to their type of response: convergent cells, those responsive to 50 °C and D R stimuli; and cells responsive only to 50 °C, or to D R stimulation. The location of the different types of CL cells is plotted in Fig. 3. We could see a h o m o g e n e o u s distribution of the convergent cells along the anteroposterior planes of the CL (planes A 4.8 to A 5.6). Also, CL cells responding to D R stimulations were found

throughout all the C L nucleus and did not have a particular distribution. C L cells responding only to 50 °C were unusual (7 of a total of 93). F u r t h e r m o r e , cortical cells also exhibit convergent responses to 50 °C and D R stimulations. PFCx cells responding to both types of stimulations were located mainly in the anterior regions (Fig. 4). In Fig. 4 we illustrate the location of different types of cortical cell responses and the location of stimulation electrode sites in the D R and adjacent structures. The m a j o r findings of this study were that: (1) cells located in CL and PFCx could r e s p o n d to both D R and 50 °C stimulations; (2) their duration responses were directly related to the duration of stimuli. Several studies have shown that the thalamic structures are capable of exhibiting responses to

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5., Fig. 3. Diagrams representing all the histological verified CL thalamic cells. CL cells were classified according to their responses to 50 °C, DR, and those responding to both types of stimulation. Drawings are taken from ref. 5.

noxious stimulation 25"29. In a detailed work Peschanski et al. 29 described the neuronal responses to noxious and non-noxious cutaneous stimuli in the p o s t e r i o r i n t r a l a m i n a r region. This includes the center- median-parafascicular complex and the centralis lateralis single unit responses. They r e p o r t e d excited or inhibited neuronal responses to noxious stimuli, but the only effect to tactile stimulation was an increase in their firing rate. In a previous work we studied non-nociceptive and nociceptive stimulation

on C L cell e v o k e d activities. We o b s e r v e d only excited cells in response to tactile stimulation and, as in the present p a p e r , we were not able to p r o v e that C L cells r e s p o n d e d with a decrease in their neuronal firing rate in response to noxious stimulation. T h e dissimilarity b e t w e e n o u r results and those r e p o r t e d by Peschanski et al. 29 could be r e l a t e d to the level and/or the type of anesthesia used, because the type of anesthesia was the only difference in the experimental design in b o t h studies,

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Fig. 4. In A, location of PFCx cells exhibiting responses to 50 °C, DR, and both types of stimulation. Note the major concentration of cells responding to both types of stimuli in plane A 9.5. In B, the points of stimulation at mesencephalic level are located. The planes in A are taken from ref. 5 and those in B from ref. 28.

The cortical participation in nociception is well documented (see refs. in the Introduction) but, to our knowledge, only in one previous work 14 were single units responding to pain stimulation reported in the rat. Our actual results confirm these cortical responses to noxious stimulation. It has been demonstrated 6"2°'21 that electrical stimulation of this cortical area (the prefrontal cortex) produces a significant reduction of nociceptive responses. These authors conclude that spinal and supraspinal mechanisms are involved in pain control and also suggest a mechanism of cortical control over pain perception. Our results show rhythmic activity in PFCx cells produced by D R stimulation that is very similar to that reported by Emmers et al. T M in the nucleus ventralis posterolateralis of the thalamic SII. These authors concluded that the rhythmic activity requires extensive summation of postsynaptic events via the feedback loops. One of them is provided by neurons of the CL and another by the P A G ~7. These two structures are also an important source of the input

to the PFCx and, thus, this is the possible origin of the rhythmic activity in cortical structures. Besides, the rhythmic activity is very frequent in CL and cortical areas 16"37. It is very interesting that all the noxious responsive cells in CL and in PFCx are also responsive to D R electrical stimulation. Moreover, the encoding capacity of CL cells to noxiousness and D R durations of the stimulation reveals the participation of these cells in pain transmission, and their relation with a serotoninergic system implicated with pain suppression like the D R 1'27. It is well established that CL cells do not have a somatotopic representation and exhibit responses to a great variety of stimuli. The simultaneous involvement of different kinds of receptor input in intralaminar neurons would then play a role in indicating a new somesthetic input without any specificity 3. Our results with noxious activated cells could be understood in the same way, but the D R input could make a special configuration of the occurrence of a new and potentially dangerous situation. Indeed, Sanders

151

et al. 35 identified n e u r o n s in the midbrain, ones

tion with activations at CL and PFCx levels. More

which increased their firing rate in the presence of

recent work 21 shows that these midbrain cells responding to noxious stimulation are also responsive to prefrontal cortical stimulation and are involved in

noxious stimuli (E-units) and those which decreased their activity in the presence of the noxious stimuli (I-units). They suggested that the E-units might represent n e u r o n s that activate the descending spinal

the analgesia process.

inhibitory pathways and 1-units might represent inhibitory i n t e r n e u r o n s which tonically inhibit the E-units, but in the presence of a noxious stimulus are themselves inhibited. Thus, the presence of a noxious stimulus would cause the removal of this tonic inhibition and thereby allow the E-units to be activated. T h e n to explain our results in this way, the midbrain system could respond to noxious stimula-

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This work was partially supported by a research grant from Consejo Nacional de Ciencia y Tecnologia CONACyT (PCEXCNA 040661 and P228CCOX880165). We thank Mr. A b e l Ortega for technical assistance, Mr. Ratil Cardoso for the illustrations and Mr. Ratil Bernal for the photography.

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