Abolition of vagino-cervical stimulation-induced analgesia by capsaicin administered to neonatal, but not adult rats

Physiology & Behavior,Vol. 44, pp. 267-272. Copyright©PergamonPress plc, 1988. Printedin the U.S.A.

0031-9384/88$3.00 + .00

Abolition of Vagino-Cervical Stimulation-Induced Analgesia by Capsaicin Administered to Neonatal, but not Adult Rats J O R G E F. R O D R I G U E Z - S I E R R A , * G E R H A R D S K O F I T S C H , t B A R R Y R. K O M I S A R U K $ A N D D A V I D M. J A C O B O W l T Z §

*Department of Anatomy, University o f Nebraska Medical Center, 42nd and Dewey Avenue, Omaha, N E 68105 tDepartment o f Zoology, University of Graz, Universitatsplatz 2, A-8010, Graz, Austr.ia ~Institute o f Animal Behavior, Rutgers-The State University, Newark, NJ 07102 §Laboratory of Clinical Science, National Institutes of Health, Building I0, Room 3D-48, Bethesda, MD 20892 R e c e i v e d 7 M a r c h 1988 RODRIGUEZ-SIERRA, J. F., G. SKOFITSCH, B. R. KOMISARUK AND D. M. JACOBOWITZ. Abolition ofvaginocervical stimulation-induced analgesia by capsaicin administered to neonatal, but not adult rats. PHYSIOL BEHAV 44(2) 267-272, 1988.--:We have previously reported that vagino-cervical mechanical stimulation (VS or probing) produces analgesia in rats. Neonatal treatment with capsaicin (CAP) has been shown to reduce the concentrations of several neuropeptides in dorsal root ganglia, spinal cord, and autonomic ganglia, via a neurotoxic effect. In the present study, we report that CAP administered in the neonatal period abolishes the analgesic effect of probing in adulthood. In addition, we report that the ability of VS to potentiate the lordosis response to manual stimulation of the flanks is abolished by neonatal CAP treatment. By contrast, rats treated as adults with CAP show the typical VS-produced effects of analgesia and potentiation of the lordosis response. Our results suggest that neonatal, but not adult, CAP treatment depletes a neuropeptide(s) that mediates the analgesia and lordosis-inducing effects of VS. Capsaicin

Pain

Vaginal stimulation

Female rats

PREVIOUS reports from one of our laboratories (B.R.K.) have shown that vagino-cervical stimulation (VS: probing against the cervix with a glass rod) inhibits behavioral responses to noxious stimulation [e.g., leg withdrawal to foot pinch (11), vocalization to tail shock (4) and tail flick to radiant heat (23)]. VS differentially suppresses the response of neurons in the somatosensory thalamus (ventrobasal complex) to cutaneous noxious stimulation but not to innocuous stimulation (12). The analgesic effect of VS has been confirmed in women; vaginal self-stimulation elevated pain detection and tolerance thresholds but did not affect tactile thresholds (30). We have demonstrated a descending brain stem-spinal cord modulatory role of monoamines (4) and a role of spinal opioid (6) and glycinergic and GABAergic systems (19,20), in VS-induced analgesia [see (30) for review]. However, the one or more primary afferent neurotransmitters that trigger this analgesia is not yet known. The pelvic nerve is a major primary afferent pathway for the vagina and cervix in rats (9,18), and bilateral transection of this nerve reduces or abolishes a variety of physiological and behavioral effects of probing (7, 10, 12, 25). The pelvic nerve (at least in cat) contains a predominance of small di-

Lordosis

Estrogen

ameter C-fibers of primary sensory neurons (9,29). Capsaicin (CAP) administered neonatally in rats permanently destroys small diameter C-fibers of primary sensory neurons (8, 15, 22, 29). Since capsaicin administered neonatally destroys Cfibers primary afferent neurons and the neuropeptides contained therein (26,29), it seems likely that administration of CAP neonatally would attenuate the effects of VS. Since CAP selectively affects unmyelinated primary sensory neurons of the C-fiber type and affects specific neuropeptides differentially depending on whether it is administered neonatally or in adulthood (22), we have also analyzed the effects of adult CAP administration of the VS effects. It has been proposed that as a result of its depleting effect on substance P, CAP administered neonatally elevates thresholds to noxious stimulation. Thus, latencies to tail flick and leg lift on the hot plate test are increased, and the number of eye wipes in response to drops of CAP solution applied to the eye are decreased, in adult rats that have received CAP neonatally. Furthermore, these rats show a reduction in substance P in the dorsal root ganglia and spinal cord (14, 22, 27). However, not all responses to noxious stimulation are attenuated by neonatal CAP. In particular,

267

RODRIGUEZ-SIERRA ET AL.

268 NEONATAL CAPSAICIN HOT PLATE TEST

[

oo z cz) ~D L~ L~

30

•

NEONATAL CAPSAICIN TAIL FLICK TEST

o

CONTROLS (n=8)

(y3

~-

F-] CONTROLS

(n=8)

o

30

z

~CAP

~CAP

(n=12)

LJ t/3

Z

Z

~- Ld z c4 kaA

>-

~ ,~

LJJ P-~a7

+

20

(_J

~

*

10

z

b

z

not tostod

BASELINE

VS

FIG. 1. Performance of rats treated neonatally with capsaicin (CAP) in the hot plate test. CAP-treated rats were not tested with vaginal stimulation (VS) because their threshold to heat was already very high (see baseline results). Details about the statistics tests used are found in the text. *p<0.05 compared to controls; '~p<0.05 compared to baseline controls.

the leg withdrawal response to foot pinch appears to remain intact and unaffected by neonatal CAP (5,14), perhaps because it utilizes A-delta fibers rather than nonmyelinated C-fibers. In the present study, we hypothesize that neonatal CAP will deplete neuropeptides in the pelvic nerve, thereby attenuating the effectiveness of this nerve in mediating the analgesia produced by VS. These neonatally CAP-treated rats are expected to be unresponsive to certain noxious stimuli (specifically, heat and chemical), but not to others (e.g., pinch). Therefore, the condition of primary interest in the present study is the foot pinch response, which is unaffected by neonatal CAP but under normal conditions is totally blocked by VS. Therefore, in the present study, we determined whether neonatal CAP treatment would prevent probing from blocking the leg withdrawal response. METHOD Female Sprague-Dawley rats (Zivic-Miller Laboratories, Allison Park, PA) were used in all experiments. Animals were housed on a 12 hr light-dark cycle and had access to food and water ad lib. Some animals received capsaicin (CAP, Sigma Chemical Co., St. Louis, MO: 50 mg/kg body weight, SC) on the second day of life, as described previously (22); controls received vehicle injections only (10% ethanol, 10% Tween 80, in normal saline). Another group of animals received CAP or vehicle at 90 to 120 days of age.

Analgesia Testing Consisted of Four Behavioral Tests Hot plate. This test consisted of placing the animal in a BeI-Art hot plate instrument (Model 475, Technilab Inc., Pequannock, NJ). The temperature was maintained at 55°C. Animals were removed from the instrument when they licked a hind paw. Latencies to lick a paw were measured by an experimenter's controlled timer. The testing was terminated if the animal did not respond by 30 seconds. Tail flick. This test consisted of placing the animal in a Plexiglas restrainer with the tail placed in a groove. The tail

BASELINE

VS

FIG. 2. Performance of rats treated neonatally with capsaicin (CAP) in the tail flick test. VS did not increase their threshold to pain in CAP-treated rats. *p <0.05 compared to controls; ap<0.05 compared to baseline controls; 'p<0.05 compared to baseline CAP-treated.

was then exposed to a high-intensity light source. A timer started at light onset and stopped when the tail moved aside from the groove and the light source. The testing was terminated if the animal did not respond by 30 seconds (in the first experiment) or by 20 seconds (in subsequent experiments). Leg withdrawal. This test consisted of pinching the rear foot of the animal with a small toothed forceps and noting the presence or absence of the leg withdrawal response. Eye wiping. This test consisted of placing a 50/xl drop of CAP (0. i mg/ml) onto the cornea. The number of times that the animal wiped its face with either front paw was counted until the animal began to explore its environment. In instances when VS was applied, it was maintained for 15 seconds and the number of wipes were counted for the remaining 45 seconds of a one-minute period. Bilateral ovariectomies were performed using halothane as an anesthetic and animals were given 7 to 10 days to recover from surgery before any testing. In the second study, estradiol benzoate (EB, 20 /xg/rat, SC; Sigma) was administered 2 days prior to testing. In the first experiment, intact neonatally CAP-treated rats and vehicle controls were given a pretest on all the analgesic tests to determine their baseline of responding. Immediately after all the baseline tests, the tests were repeated in combination with VS. In the eye wiping test, the left eye was tested during the baseline test and the right eye was tested during the VS test. Lordosis responsiveness was tested at the onset of the VS testing period, as this response wanes with repeated stimulation (21). After all the tests were completed, all animals were tested again with and without VS. In the second experiment, intact adult rats were given a pretest on all the analgesic tests described above. Immediately after receiving the baseline tests, the tests were repeated with half of the animals receiving VS and the other half receiving no VS. On the next day, all animals were anesthetized with halothane to prevent the occurrence of writhing that occurs after CAP treatment in adult animals. Half of the animals were administered CAP (50 mg/kg, SC) and the other half of the rats received the vehicle injection. Five days after CAP or vehicle injections, all animals received all tests again, without VS. After these baseline tests,

CAPSAICIN: A D U L T VS. N E O N A T E S

269 NEONATAL CAPSAICIN-ESTROGEN TREATED

TABLE 1 NEONATAL TREATMENTWITH CAPSAICIN. EFFECTS ON NOCICEPTIVERESPONSES IN THE FEMALE RAT

HOT PLATE TEST (j-)

Behavior: Eye Wipes (Mean Frequency +_ S.E.M.) Group Controls Capsaicin

(n) (8) (12)

Baseline 28.7 _+ 1.5 0

VS 13.1 +_ 2.8* 0

c= Z

30

o i,i

[]

CDNTROLS (n=3)

~

CAP

-X-

o

=

z

~: >- ,,~

20

Behavior: Leg Withdrawal to Pinch (Percent Responding) Controls Capsaicin

(8) (12)

100% 100%

0%* 100%

Behavior: Lordosis to Manual Stimulation (Percent Responding) Controls Capsaicin

(8) (12)

0% 0%

< -J

v

10

t.J

100%* 0%

BASELINE

VS

*p<0.05.

NEONATAL CAPSAICIN - ESTROGEN TREATED

FIG. 3. Performance of ovariectomized rats treated neonatally with capsaicin (CAP) and given estradiol benzoate (EB) two days before being evaluated by the hot plate test. *p <0.05 compared to controls; ap<0.05 compared to baseline controls. CAP-treated animals did not differ due to testing with VS.

TAIL FLICK TEST

RESULTS

03 t"n Z CD U c.r) Z

I [ ] CONTROLS (n=3) 20

EXPERIMENT

cAP (~=6)

1. E F F E C T S

OF NEONATAL

CAPSA1CIN

TREATMENT

Baseline Responses ~--

>- ~ u z ~ ~--

[]

o

+

10

-..J

BASELINE

VS

FIG. 4. Performance of ovariectomized rats treated neonatally with capsaicin (CAP) and given estradiol benzoate (EB) two days before being evaluated by the tail flick test. *p <0.05 compared to controls; ap<0.05 compared to baseline controls. CAP-treated animals did not differ due to testing with VS.

Pain thresholds. The neonatally CAP-treated rats showed significant longer latencies on the tail flick and hot plate tests than the controls (Figs. 1 and 2). Furthermore, they showed no responses to the CAP drops in the eye, whereas all the controls did respond to this noxious stimulus (Table 1). By contrast, the leg withdrawal response was unaffected by neonatal CAP administration (i.e., the CAP and control groups both showed a normal leg withdrawal response) (Table 1). Thus, the neonatal CAP reduced or abolished responsiveness to noxious thermal or chemical stimulation but not to noxious mechanical stimulation. Lordosis. Lordosis was not elicited in the baseline tests in response to palpation of the flanks in the experimental or control rats (Table 1). Responses to VS

half of the animals in each group (CAP or vehicle) were retested without VS and the other half of the group was tested with VS. The next day the groups were reversed, so that after CAP, all animals were tested twice without VS and once without VS. In the third experiment, rats were bilaterally ovariectomized and allowed to recover for one week prior to EB treatment. Two days after EB, all animals were pretested without VS and then tested again, half of the animals receiving VS and the other half receiving no VS. Two days after the test, haft of the animals received CAP (50 mg/kg, SC) while anesthetized with halothane. Three days after CAP or vehicle all rats received EB. Two days after EB, all animals were tested without VS. After this baseline test, half of the animals in each group (CAP and vehicle) were retested without VS and the other half was tested with VS. The next day the group conditions were reversed, so that after CAP, all the animals were tested twice without VS and once with VS.

Pain thresholds. Control rats (no neonatal CAP treatment). As expected, in the control rats VS significantly elevated latencies on the tail flick and hot plate tests (Figs. 1 and 2) and significantly reduced the number of eye wipes to CAP applied to the eye (Table 1). Furthermore, as expected in these controls, VS abolished the leg withdrawal response to foot pinch (Table 1). Neonatal CAP-treated rats. In marked contrast to the control rats, in these CAP-treated rats, VS was completely ineffective in blocking leg withdrawal to foot pinch (Table 1). In other words, the CAP-treated rats continued showing leg withdrawal responses to foot pinch when VS was applied, whereas in the control rats, VS abolished the leg withdrawal response to foot pinch. Since the neonatal CAP-treated rats were essentially unresponsive on the other tests (i.e., hot plate, tail flick latency and CAP in the eye), they were not tested systematically with VS on those tests, since a blockage of responsiveness would not be observable.

270

R O D R I G U E Z - S I E R R A E T AL.

TABLE 2 NEONATAL TREATMENT WITH CAPSAICIN. EFFECTS ON ESTROGEN ON NOCICEPTIVE RESPONSES IN THE FEMALE RAT

HOT PLATE TEST

Behavior: Eye Wipes (Mean Frequency + S.E.M.) Group Controls Capsaicin

(n) (3) (12)

Baseline 32.0 _+ 1.5 0.3 ± 0.3

VS 9.3 ± 2.8 0

Behavior: Leg Withdrawal to Pinch (Percent Responding) Controls Capsaicin

(8) (12)

10~Vc~ 10lY)~

0%* 100~/~

Behavior: Lordosis to Manual Stimulation (Percent Responding) Controls Capsaicin

(8) (12)

100% 100%

w

Lf3

z

F []

•

CONTROLS

b

o

20

>- LZ

~L _1

]S

-7

BASELINE 1

BASELINE 2

VS

EXPERIMENT 3. EFFECTS OF ADULT CAPSAtCIN TREATMENT

~

Baseline Responses

r 3

+

j

(n=10)

=

(n=iO)

>- LA

~

z

•

CONTROLS

"~C

FIG. 5. Performance of ovariectomized rats treated as adults with capsaicin (CAP) in the hot plate test. CAP-treated animals had higher thresholds to pain in both baseline tests. VS increased the threshold of the control group and CAP-treated significantly. *p<0.05 compared to controls; ~?~<0.05 compared to baseline tests.

AOULT CAPBAICIN TAIL FLICK TEST

20

[ 30

z c~ o L~ C~

10ff~. 83%

*p <0.05.

z cz~ ~D

AOULT EAPSAIEIN

10

_1

0

BASELINE ]

BASELINE 2

VB

FIG. 6. Performance of ovariectomized rats treated as adults with capsaicin (CAP) in the tail flick test. Note that although CAP-treated animals had higher thresholds to pain in both baseline tests, VS increased the threshold of both groups significantly. *p<0.05 compared to controls; ap<0.05 compared to baseline tests.

Pain thresholds. Tail flick latency and hot plate response latency were both significantly longer in the rats treated in adulthood with C A P than in the controls (Figs. 5 and 6). The n u m b e r of eye wipes in response to C A P placed into the eye was significantly reduced in the adult CAP-treated rats compared to the controls (Table 3). The leg withdrawal response to foot pinch did not differ b e t w e e n the C A P and the control groups. Thus, on all baseline tests, the effect of C A P treatm e n t in adulthood was similar to that o f neonatal C A P treatment. Lordosis. Since estrogen was not administered, none of the rats showed lordosis in response to flank palpation only, the same results as in the neonatal C A P experiment (Table 3).

Response to VS Lordosis. In the control (non-CAP) rats, VS plus flank stimulation elicited lordosis. By contrast, in the neonatally C A P - t r e a t e d rats, VS plus flank stimulation failed to elicit lordosis (Table 1). Lordosis could not be elicited by either VSonly or flank stimulaton-only in either group. EXPERIMENT 2. EFFECTS OF ESTROGEN TREATMENT AFTER CAPSAICIN Administration of estradiol benzoate after o v a r i e c t o m y yielded results essentially the same as in E x p e r i m e n t 1 (Figs. 3 and 4 and Table 2), e x c e p t for the lordosis tests, as follows. W h e n estrogen was administered to the rats, all s h o w e d lordosis to flank palpation, controls as well as neonatal CAPtreated. VS p r o d u c e d no greater effect in any o f the rats. Thus, neonatal C A P did not block the manifestation of lordosis per se, but it did block the normal ability o f VS to facilitate the lordosis response to flank palpation.

Pain thresholds. Control animals. VS significantly increased the tail flick latencies and hot plate response latencies o v e r pre-VS control levels, and significantly d e c r e a s e d the n u m b e r of eye wipes from pre-VS levels (Figs. 5 and 6 and Table 3). Leg withdrawal responses were abolished by VS (Table 3). Adult CAP-treated animals. In marked contrast to the effects seen with neonatal C A P treatment, the VS effect was not blocked on any measure in the adult C A P - t r e a t e d group. That is, VS significantly increased tail flick latencies and hot plate response latencies in the CAP-treated as well as in the control groups (Figs. 5 and 6). VS abolished both the eye wipe response after C A P into the eye and the leg withdrawal response to foot pinch in both groups (Table 3). Lordosis. In the control as well as the adult CAP-treated rats, VS plus flank palpation induced lordosis in all individuals (Table 3).

C A P S A I C I N : A D U L T VS. N E O N A T E

271

TABLE 3 ADULT TREATMENTWITH CAPSAICIN. EFFECTS ON NOCICEPTIVERESPONSES IN THE FEMALE RAT Behavior: Eye Wipes (Mean Frequency -4- S.E.M.) Groups Controls Capsaicin

(n) (10) (9)

Baseline 1 30.8 _+ 1.3 1.0 _+ 1.0

Baseline 2 31.8 -+ 1.3 0

VS 9.0 _+ 2.2 0

Behavior: Leg Withdrawal to Pinch (Percent Responding) Controls Capsaicin

(10) (9)

100~ 100%

100% 100%

0% 0%

Behavior: Lordosis to Manual Stimulation (Percent Responding) Controls Capsaicin

(10) (9)

0% 0%

0% 0%

100% 100%

*p <0.05.

DISCUSSION Our results clearly demonstrate that after neonatal CAP treatment, female rats are refractory to VS; they show neither the analgesic nor the lordosis-facilitatory effects of VS. By contrast, after CAP treatment in adulthood, female rats remain responsive to VS. The failure of CAP to block VS-produced analgesia when administered to adult rats confirms the findings of Nance et al. (16), who administered CAP intrathecally. However, our results differ from those authors in that CAP treatment in adulthood attenuated VSproduced lordosis facilitation in their study, but it did not do so in ours. The difference may be due at least in part, to the systemic vs. intrathecal administration. It is also possible that a quantitative depletion of (a) critical neuroactive transmitter(s) could be reflected as a qualitative difference effect. Thus, if analgesia in response to VS can be produced with less transmitter than lordosis, a subtotal depletion of transmitter related to route of CAP and/or age of administration, could result in a differential blockage of the lordosis but not the analgesia response to VS. Both treatments (neonatal and

adult CAP) result in the previously described increased thresholds to noxious stimuli (2, 8, 13). Since CAP treatment appears to deplete substance P equally in neonates and adult female rats (13,22), this depletion may be the basis for the increased thresholds to noxious stimuli. In addition, on the basis that the concentration of substance P (SP) in the vagina, cervix and uterus is reduced by neonatal administration of CAP (27) and that certain neuroendocrine reflexes (e.g., pseudopregnancy) that are blocked by surgical transection of the pelvic nerve (3) which produces sensory innervation of the vagina and cervix (18), are also blocked by neonatal CAP administration, it has been suggested that SP mediates vaginal afferent activity (28). However, other substance(s) could also mediate the effects of VS. Since CAP depletes a number of neuropeptides other than SP in spinal cord, dorsal root ganglia, and autonomic ganglia, it is possible that one or more of these neuropeptides is the neuromediator for the VS effects. Three other neuropeptides have been shown to be affected by CAP; vasoactive intestinal peptide (VIP), somatostatin (SS), and corticotropin releasing factor (CRF) (17,22). All three of these peptides have been localized in sensory regions of the spinal cord (22,27). One of these neuropeptides, VIP, has been shown to be differentially affected by neonatal vs. adult CAP treatment: it is depleted by CAP in the adult animal to a lesser extent than in neonatally-treated rats (22). Furthermore, VIP is present in the pelvic nerve [colocalized with SP (9)] and is released into spinal cord superfusates after electrical stimulation of the nerve in cats (1). In addition, VIP has been shown to produce analgesia when injected into the midbrain periaqueductal gray (24) or into the intrathecal space of the spinal cord. The above findings are consistent with a role of VIP in mediating the analgesic and possibly other effects of VS.

ACKNOWLEDGEMENTS This work is partially supported by a Faculty Development Fellowship from the University of Nebraska Medical Center and by the NIH (HD-13219 to J.F.R.-S.). This is contribution No. 476 from the Institute of Animal Behavior.

REFERENCES 1. Blank, M. A.; Anand, P.; Lumb, B. M.; Morrison, J. F.; Bloom, S. R. Release of vasoactive intestinal polypeptide-like immunoreactivity (VIP) from cat urinary bladder and sacral spinal cord during pelvic nerve stimulation. Dig. Dis. Sci. 29:11 ; 1984. 2. Bodnar, R. J.; Simone, D. A.; Kordower, J. H.; Kirchsgessner, A. L.; Nilaver, G. Capsaicin treatment and stress-induced analgesia. Pharmacol. Biochem. Behav. 18:65-71; 1983. 3. Carlson, R. R.; DeFeo, V. J. Role of the pelvic nerve vs. the abdominal sympathetic nerves in the reproductive function of the female rat. Endocrinology 77:1014-1022; 1965. 4. Crowley, W. R.; Rodriguez-Sierra, J. F.; Komisaruk, B. R. Monoaminergic mediation of the antinociceptive effect of vaginal stimulation in rats. Brain Res. 137:67-84; 1977. 5. Doucette, R.; Threriault, E.; Diamond, J. Regionally selective elimination of cutaneous thermal nociception in rats by neonatal capsaicin. J. Comp. Neurol. 261:583-591; 1987. 6. Faris, P. L.; Komisaruk, B. R.; Watkins, L. R.; Mayer, D. J. Evidence for the neuropeptide cholecystokinin as an antagonist of opiate analgesia. Science 219:310-312; 1983.

7. Heller, S. B.; Gintzler, A. R.; Stracher, A.; Komisaruk, B. R. Vaginal stimulation-produced analgesia is prolonged by leupeptin, a protease inhibitory. Soc. Neurosci. Abstr. 11:124; 1985. 8. Jancso, N.; Jancso-Gabor, A.; Szolcsanyi, J. Direct evidence tbr neurogenic inflammation and its prevention by denervation and by treatment with capsaicin. Br. J. Pharmacol. Ther. 31:138-151; 1967. 9. Kawatami, M.; Nagel, J.; DeGroat, W. C. Identification of neuropeptides in pelvic and pudendal nerve afferent pathways to the sacral spinal cord of the cat. J. Comp. Neurol. 249:117132; 1986. 10. Komisaruk, B. R.; Banas, C.; Heller, S. B.; Whipple, B.; Barbato, F.; Jordan, F. Evidence that vasoactive intestinal peptide administered directly to the spinal cord produces analgesia in rats. Soc. Neurosci. Abstr. 13:1311; 1987. 11. Komisaruk, B. R.; Larsson, K. Suppression of a spinal and a cranial nerve reflex by vaginal or rectal probing in rats. Brain Res. 35:231-235; 1971. 12. Komisaruk, B. R.; Wallman, J. Antinociceptive effects of vaginal stimulation in rats: neurophysiological and behavioral studies. Brain Res. 137:85-107; 1977.

272 13. Lembeck, F. Zur Frage der Zentralen Ubertragung afferenter Impulse. III. Mitteilung Das Vorkomen und die Bedeutung der Substanz P in den dorzalen Wurzeln des Ruckenmarks. Naunyn Schmiedebergs Arch. Exp. Pathol. Pharmacol. 219:197-213; 1953. 219:197-213; 1953. 14. Lembeck, F.; Skofitsch, G. Visceral pain reflex after pretreatment with capsaicin and morphine. Arch. Pharmacol. 321:116122; 1982. 15. Nagy, J. 1. Capsaicin: a chemical probe for sensory neuron mechanisms. In: Iversen, L. ; Iversen, S. D. ; Snyder, S. H., eds. Handbook of psychopharmacology. New York: Plenum Press; 1982:185-235. 16. Nance, D. M.; King, T. R.; Nance, P. W. Neuroendocrine and behavioral effects of intrathecal capsaicin in adult female rats. Brain Res. Bull. 18:109-114; 1987. 17. Ottensen, B.; Larsen, J. J.; Staun-Olsen, P.; Gammeltoft, S.; Fahrenkrug, J. Influence of pregnancy and sex steroids on concentration, motor effect and receptor binding of VIP in rabbit female genital tract. Regul. Pept. 11:83--92; 1985. 18. Peters, L. C.; Kristal, M. B.; Komisaruk, B. R. Sensory innervation of the external and internal genitalia of the female rat. Brain Res. 408:199-204; 1987. 19. Roberts, L. A.; Beyer, C.; Komisaruk, B. R. Strychnine antagonizes vaginal stimulation-produced analgesia at the spinal cord. Life Sci. 36:2017-2023; 1985. 20. Roberts, L. A.; Beyer, C.; Komisaruk, B. R. Nociceptive responses to altered Gabaergic activity at the spinal cord. Life Sci. 39:1667-1674; 1986.

RODRIGUEZ-SIERRA

ET AL.

21. Rodriguez-Sierra, J. F.; Komisaruk, B. R. Estrogen accelerates the recovery of the lordosis response after its exhaustion induced by cervical probing. Horm. Behav. 17:302-307; 1985. 22. Skofitsch, G.; Zamir, N.; Heike, C. J. ; Savitt, J. M.; Jacobowtiz, D. M. Corticotropin releasing factor-like immunoreactivity in sensory ganglia and capsaicin sensitive neurons of the rat central nervous system: colocalization with other neuropeptides. Peptides 6:307-318; 1985. 23. Steinman, J. L.; Komisaruk, B. R.; Yaksh, T. L.; Tyce, G. M. Spinal cord monoamines modulate the antinociceptive effects of vaginal stimulation in rats. Pain 16:155-166; 1983. 24. Sullivan. T. L.; Pert, A. Analgesic activity of non-opiate neuropeptides following injections into the rat periaqueductal gray matter. Soc. Neurosci. Abstr. 7:504; 1981. 25. Szechtman, H. ; Adler, N. T. ; Komisaruk, B. R. Mating induces pupillary dilatation in female rats: Role of pelvic nerve. Physiol. Behav. 35:295-301; 1985. 26. Szolcsanyi, J. Capsaicin and nociception. Acta Physiol. Hung. 69:323-332; 1987. 27. Traurig, H.; Saria, A.; Lembeck, F. Substance P in primary afferent neurons of the female rat reproductive system. Arch. Pharmacol. 326:343-346; 1984. 28. Traurig, H.; Saria, A.; Lembeck, F. The effects of neonatal capsaicin treatment on growth and subsequent reproductive function in the rat. Arch. Pharmacol. 327:254-259; 1984. 29. Wall, P. D. The central consequences of the application of capsaicin to one peripheral nerve in adult rat. Acta Physiol. Hung. 69:275-286; 1987. 30. Whipple, B.; Komisaruk, B. R. Elevation of pain threshold by vaginal stimulation in women. Pain 21:357-367; 1985.