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Keywords:

  • erectile dysfunction;
  • animal model;
  • cavernous nerve

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

OBJECTIVE

To develop a rat model of erectile dysfunction (ED) after cavernous nerve injury.

MATERIALS AND METHODS

Given the great similarity between the anatomical structure of the cavernous nerve in rats and humans, 24 rats underwent dissections and the cavernous nerves were identified with the aid of an operating microscope. Then the rats were randomized into two groups: sham-operated controls and a bilateral cavernous nerve section group. At 3 months after surgery, the rats were evaluated for their response to an apomorphine challenge.

RESULTS

The erectile response after an apomorphine challenge was normal in all the control rats, while there were no erections in the bilateral injured group.

CONCLUSION

The rat major autonomic ganglion and its cavernous nerve can be identified with the aid of a microscope. Rats are inexpensive and easy to handle, thus a good animal for developing an ED model of cavernous nerve injury. In the present study, the rats with cavernous nerve injury lost erectile capacity in a reliable and reproducible fashion. Because of the great similarity between the cavernous nerve of rats and humans, one may consider this technique as a reliable experimental model for studying ED after radical prostatectomy.


Abbreviations
ED

erectile dysfunction

RP

radical prostatectomy

PVN

paraventricular nucleus.

INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

Erectile dysfunction (ED) is recognized as a medical condition, presenting multifactorial and complex aetiology related to the ageing of the population and plurimetabolic diseases. Radical prostatectomy (RP) is the main technique used for curative treatment of localized prostate carcinoma [1,2]. However, ED is a major complication of this surgery, generally caused by injury of the cavernous nerve given its close proximity to the prostate [3]. Traction, laceration and compression are the most common types of peripheral nerve injury potentially leading to ED in these patients [4]. Based upon these facts there has been a resurgence of interest in basic research of the erectile process under normal and abnormal conditions.

Many animal models of cavernous nerve injury, using different species such as dogs and monkeys, have been used in previous studies of this kind due to the facility of identification of the cavernous nerve, bigger than the rats' similar plexus, which is too small to be identified with the naked eye. The purchase and maintenance costs of these large animals for medical research has become a problem. Because of the great similarity between the cavernous nerve of rats and humans, in addition to cost-effectiveness and easy handling, we developed a rat model of ED, identifying the rat cavernous nerve and lesioning this nerve.

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

In all, 24 male Wistar-EPM rats (3 months old, 300–350 g) were randomly divided into two equal groups. One group (uninjured controls) had a sham operation, while the remaining group had bilateral cavernous nerve injury. All institutional and national guidelines for animal safety and comfort were strictly followed and surgery was done with local ethical approval and is consistent with global ethical requirements.

The surgery was done under ketamine and xylazine anaesthesia. Isothermia was maintained at 37 °C by placing the rats on a heating pad. A lower abdominal midline incision was made from the symphysis pubis to mid-abdomen. The testes were retracted, their gubernacula were divided and they were packed into the upper abdomen. An operating microscope aided dissection. The cavernous nerve exits the major pelvic ganglion in the groove between the urethra and rectum, dividing under the symphysis and finally innervating the bulbous urethra and corpora cavernosa. There were also were numerous fine nerve fibres coursing from the major pelvic ganglion in all directions toward the pelvic viscera. The endopelvic fascia overlying the cavernous nerve was incised and after periprostatic dissection, the nerve and the major pelvic ganglion were identified posterolaterally on either side of the prostate (Fig. 1). The uninjured controls had no further manipulation. In the remaining group the cavernous nerves were isolated and then injured by sharply excising a segment creating a 3 mm gap. Collateral fibres to the cavernous nerves were also carefully dissected. The procedure was repeated on the contralateral side.

image

Figure 1. The cavernous nerve after periprostatic dissection and opening of the endopelvic fascia.

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At 3 months after the surgical procedure both groups were given the apomorphine test. Apomorphine solution (0.05 mg/kg) was injected in the loose skin of the back of the neck to induce an erection, based upon the fact that microinjection of apomorphine (a dopamine agonist) into the paraventricular nucleus of the hypothalamus (PVN) induced penile erection in rats and was identical to that induced by the systemic administration of the drug, suggesting that the PVN is the brain area where dopamine agonists act to induce penile erection [5]. After apomorphine administration each rat was observed for 30 min, especially for penile erections. An erection was only counted when the emergence of an engorged glans penis and distal shaft was noted (Fig. 2).

image

Figure 2. Erection after apomorphine test injection.

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RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

As shown in Fig. 3 the rat major pelvic ganglion lies on either side of the dorsolateral lobes of the prostate. It receives innervation from the sacral parasympathetic nuclei via the pelvic nerve and from thoracolumbar outflow via the hypogastric nerve. The largest efferent nerve runs along the lateral aspect of the urethra without branching. This nerve is termed the cavernous nerve.

image

Figure 3. Anatomical and line drawing of the cavernous nerve and its location.

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The apomorphine test had an effect a few minutes after s.c. injection and it lasted for ≈30 min. During this period, all rats were more active, with hair standing, stretching and yawning. All the rats in the control group (uninjured nerves) responded to the apomorphine test with erections for several minutes. In the bilateral cavernous nerve injury group there were no erections in any of the rats. When this group was compared with the control rats there was a complete loss of erectile capacity for a period as long as 3 months after surgery.

DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

RP for prostate cancer is a widely utilized curative treatment for localized prostate carcinoma [2] but the sequelae, such as ED and urinary incontinence, have profound effects on quality of life. Walsh et al.[6] popularized nerve-sparing RP, which decreases the rate of ED after RP but many patients at risk of positive margins by virtue of stage (≥T2b), grade (≥Gleason 7) or PSA level (>15 ng/mL) may not be candidates for a nerve-sparing procedure [7]. The cavernous nerve has to be resected bilaterally or unilaterally in the process of radically removing the cancer. Like other peripheral nerves the cavernous nerves have the intrinsic capacity to regenerate fully and restore function, at least in limited nerve lesions. However, after the nerve is transected completely the guidance structure allowing orientated regrowth and proper target reinnervation is missing [8]. Transection of the fibre results in distal fragmentation of axon and myelin sheaths [9].

Although the precise anatomy may differ from species to species, the overall framework of rat pelvic autonomic innervation is remarkably similar to that of humans and other animals. In mammals, the main constitution is the pelvic plexus that is formed by the joining of the thoracolumbar sympathetic fibres via the hypogastric nerve and sacral parasympathetic fibres via the pelvic nerve. The largest efferent branch is the cavernous nerve. In rats, the major pelvic ganglion and its cavernous nerve can be easily identified with the aid of a surgical microscope. With reference to the basic anatomy of the cavernous nerve [10,11], the major pelvic ganglion were exposed in the posterolateral surface of the dorsal lobes of the prostate. Based on the ganglion, its inflows (hypogastric nerve and pelvic nerve) and outflow (cavernous nerve) were all identified.

At 3 months after cavernous nerve injury, the rat model was evaluated using the apomorphine test. It is well documented that a low dose of apomorphine causes penile erection, stretching and yawning syndrome in rats [5]. This action is mediated by dopamine receptors in the PVN of the hypothalamus. In the present study, all rats after apomorphine injection were more active, with hair standing, stretching and yawning. All rats in the control group had recorded penile erections. Conversely, for the cavernous nerve injury group, there were no erections, suggesting that bilateral cavernous nerve injury can lead to ED after injury. The apomorphine test evaluates erectile response directly, but it cannot show alterations in haemodynamics in the corpus cavernous tissue. The present bilateral cavernous nerve injury proved to be a simple and reproducible method resulting in the loss of penile erection. We could easily identify the rat cavernous nerve, induce injury and develop a rat model of ED caused by cavernous nerve injury. This rat model could be widely used as rats are inexpensive, readily available and the cavernous nerve can be easily identified.

In conclusion, the rat major ganglion and its cavernous nerve can be identified with the aid of a microscope. Rats are inexpensive and easy to handle, thus making a very good animal for developing an ED model caused by cavernous nerve injury. The present study showed that rats in the group with cavernous nerve injury consistently lost erectile capacity, thus producing a reliable model of ED caused by cavernous nerve injury. Because of the of the great similarity between the cavernous nerves of rats and humans in addition to ready availability, rats may constitute an appropriate experimental model for studying ED after RP.

ACKNOWLEDGEMENTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES

Financial support provided by Institutos do Milenio de Bioengenharia Tecidual, CNPq (Brazil).

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. ACKNOWLEDGEMENTS
  8. CONFLICT OF INTEREST
  9. REFERENCES