World without Cats (32 page)

On the appointed day, Vera and Kal drove to the institute with the materials and equipment they needed for semen collection. Jane remained at the clinic in case she was needed to bring anything they’d forgotten.

When they arrived at Noah’s office, Vera declared, “It’s showtime.”

“What happens now?” Noah asked.

“Today, we do three procedures. First we’ll inject three of the females with gonadotropin to stimulate superovulation. That way, we should have plenty of oocytes ready to harvest tomorrow. Second, we’ll inject four other females—the ones that will receive the embryos later—with FSH to induce estrus.”

“What’s FSH, and why do the mother cats have to be in estrus? They’re not going to be mating, are they?”

Vera explained, “FSH stands for follicle-stimulating hormone. We shoot it into muscle tissue every day, until the queen shows signs of estrus. This should take three or four days. The reason we want the donor queens in estrus is that the embryo transplants are only likely to take if the recipient cat is in the same physiological state as the mother cat would be if the ovum had been fertilized naturally.”

“Whew! I had no idea there was so much to this.”

“Yeah, it’s almost as complicated as molecular biology.” Vera smiled wryly. “Anyway, once the mother-to-be is in estrus, we inject gonadotropin, just as we’ll do with the egg donors. Also, today, we plan to collect semen from the toms.”

Noah wondered, “How do you do that? Masturbation?”

“No. That’s one way—some vets use an artificial vagina—but it’s much easier to use electroejaculation.”

“Aaghh! Doesn’t sound humane.”

Vera grinned. “Actually, it is. The male is anaesthetized and has no awareness of the procedure. When he wakes up, he’s normal and in no pain. Who knows, maybe he has an erotic dream during the process.”

Noah chuckled. “Is there anything I can do to help?” he asked.

“Not with the semen collection. We’ll extract the oocytes from three of the queens tomorrow. After we do the IVF, I’ll need your magic bullet prep. You can thaw it and hold it on ice until we’re ready.” They were now referring to the FeLV vector with its attached FHF genes as the “magic bullet,” an expression popularized over a century earlier by the German chemist Paul Ehrlich to refer to chemicals that fight microbial infections without harming the patients.

“What’s IVF?” asked Noah.

“In-vitro fertilization.”

“Oh, right. You guys use as much jargon molecular biologists, don’t you? I’ve got a dozen vials of the vector in the ultra-cold freezer. I’ll thaw a couple of them as soon as I get in tomorrow morning.”

“Fine. I’ll have Jane and Kal with me to help with the surgery. Jane will act as surgical nurse and Kal as an assistant surgeon.” She frowned. “Sounds pretentious, doesn’t it?”

Noah smiled. “I think you’re entitled to any pretentions you fancy.”

She squeezed his hand. “One thing you can do,” she said, “is help Doris look after Lilith. I’m going to be spending more time here for the next few days than I have recently.”

“No problem. I can run home in the middle of the day.”

Vera made use of the shower adjacent to the lab, washing down thoroughly with hexachlorophene soap. Kal did the same. The two vets donned sterile surgical scrubs, complete with facemasks, hoods, and paper booties over their shoes. They scrubbed their arms a second time with surgical soap and Betadine. No way were they going to introduce any FHF into the lab. Before entering, Vera switched off the UV lamps.

“Okay, that’s it,” Kal remarked, after he’d injected the three females with gonadotropin. “Let’s go collect the semen. Romeo’s first, right?”

In keeping with the tradition set in Noah’s lab, the thirteen feral cats had been named after Shakespearian characters. After several days in captivity, the first donor, Romeo, had become somewhat conditioned to his cage and to the lab environment. Kal, with the help of a pinch of catnip, met little resistance when he opened the sliding door and reached in. With a small electric clipper, he shaved Romeo’s right forepaw to expose the cubital vein, while Vera readied a syringe containing ketamine mixed with diazepam. While Kal held the tom steady, Vera injected the mix.

“He’s under,” Vera said. Kal took the teflon-coated electrode and inserted it in the cat’s rectum. He switched on the power supply and applied a succession of rapid-fire stimuli. Vera collected the resulting semen in a small vial containing a holding buffer. When they were done, Kal gently placed Romeo back in his cage. The vets collected specimens from two other toms, Polonius and Lear, in the same manner. The semen would be used the next day, so Vera decided simply to keep the vials unfrozen in an ice bath overnight.

Later, when Kal examined the first of the semen samples with Noah’s microscope, he raised a fist in the air and shouted, “Yes! Good sperm count with excellent motility.”

Vera let out a lungful of breath she’d unconsciously held in. “Ah, good,” she sighed.

The other two specimens showed favorable sperm counts also. Before they left for the day, Vera and Kal removed the food dishes from the two gonadotropin-treated females. Tomorrow, the queens would be egg donors.

Vera, Kal, and Jane arrived at six the next morning, carrying several large cases of surgical supplies—instruments, a small tank containing compressed carbon-dioxide gas, anesthetics, buffers, and the like. As soon as she had showered and put on sterile garments, Vera took the semen samples out of the ice bath to warm them. They would sit at room temperature for several hours to allow capacitation of the spermatozoa. She then removed the water dishes from the two females that were to serve as egg donors.

Kal and Jane soon joined Vera and prepared for the surgery. “All right, team,” Vera pronounced officiously, “let’s review. The procedure is to extract egg cells from the donor queens, fertilize the ova
in vitro
with semen collected yesterday, and, after fertilization occurs, to inject the bullet vector into the nuclei of the cells. Kal, continue.”

“Testing me, are you? Okay, then. We’ll incubate the fertilized eggs for four or five days, until the cells divide a few times. Finally, we’ll transplant the embryos into the four recipient females. With luck, they’ll develop into FHF-resistant kittens.”

“Good, Kal.” Vera shook his hand. She hugged him, and Jane joined. They remained in a tri-part huddle for almost a minute.

After several hours, everything was ready. “I think we should eat something before we start,” Vera suggested. None of them had eaten breakfast, and it was now approaching nine o’clock.

After removing their special clothing, they took a quick meal at the campus cafeteria, with Noah along to offer what encouragement he could. When they returned, the three again went through the tedious process of showering, disinfecting, and donning the scrubs.

While Noah watched through a window, Vera anaesthetized Ophelia, the first of the queens to donate her eggs. When she was sure that the cat was unconscious, Vera positioned her on the surgical table.

“Tie her limbs, will you, Kal?”

He quickly tied Ophelia’s four legs to hooks at the sides of the table. Jane grasped electric shears and shaved the skin of the cat’s lower abdomen, after which Vera swabbed the area with Betadine and surgical soap. Kal then draped sterile cloths over most of the animal, leaving only a small rectangle of skin exposed.

“Okay, Kal, inject the carbon dioxide.”

Kal picked up a sterile needle attached to a long, flexible length of tubing that extended from the small tank. He inserted it through the exposed skin of the cat into the abdomen. He nodded to Jane, who twisted the valve on the tank through a small arc. The trio could see the abdomen begin to expand.

“It’s like blowing up a balloon,” Jane remarked. “Why are we doing this? I forgot.”

“It makes it much easier to work inside. If all the organs were sitting on top of one another, it would be difficult to get around with these delicate instruments.”

When all was ready, Vera made small cuts with a scalpel, first through the dermis, and then through the abdominal wall. With a grabber, she pulled the cat’s left ovary toward the opening and handed the instrument over to Jane as she said, “I want you to hold this as steady as you can.”

“Right.”

Vera nodded. “Kal, would you position the laparoscope, please?”

She took hold of the probe with her right hand and the aspirating needle with her left. Jane held the collecting tube with her other hand.

Vera checked the computer screen as she moved the probe. “Hah!” she announced after a few seconds. “Several oocytes to choose from. Suction, please.” Kal turned on the aspirator, and Vera maneuvered the needle over the ovary, where she could see the oocytes. Each time she lowered the needle, she heard a faint
sshloop
as small bits of ovary were sucked into the collecting tube.

“All right, that’s three so far. Let’s get some from the other side.” Vera took the grabber from Jane and allowed the ovary to settle back into its natural position. “Now pull up the right ovary,” she said, handing the grabber back to Jane. Vera then carefully guided the probe to the organ and harvested four more oocytes from the right side.

When they were done with Ophelia, Kal carefully closed the small incision. Gently, he carried Ophelia to her cage to recover. Vera waved to Noah through the window and held up her thumb in an okay sign. Noah pointed to his watch and indicated that he had to get to work. By mid-afternoon, the three surgeons had finished collecting oocytes from the other two females, Portia and Goneril.

“There,” said Vera, pointing to the three small plastic culture dishes, “rest twenty-four microscopic cells that might provide the only hope for the survival of the domestic cat.”

Vera and Kal now proceeded to carry out in vitro fertilization with feline semen collected the previous day. Vera sat down at the stereomicroscope and, with the help of the micromanipulator, carefully teased oocytes away from the tissue, transferring them to dishes containing buffered nutrients and antibiotics to inhibit bacteria and molds. For the next five hours, she and Kal took turns at the stereomicroscope, examining the dishes for signs of successful IVF.

Near the end of the first hour, Kal shouted, “Got one!”

“Let’s see.” Vera peered through the oculars and immediately spotted the thickened outer membrane on one of the oocytes, characteristic of fertilization. She could barely make out the two pronuclei—one from the egg and one from the sperm—until she cranked up the magnification. “Well, here we go. One small step for a vet, one giant leap for catkind … or so we hope.”

As Kal and Jane looked on, Vera removed the fertilized egg to its own tiny transparent plastic dish. She loaded a small amount of the FeLV-FHF vector constructed by Noah and Gary into an extremely thin glass pipette attached to the micromanipulator. Abruptly, Vera stood up and, eyes on the floor, paced around the room.

“What’s the matter?” asked Kal.

“I’m just nervous, I guess. There’s so much riding on this … we can’t fail this time. We just can’t.”

Jane reached for Vera’s latex-gloved hands and took them in her own. She said nothing, but looked into Vera’s eyes, an expression of empathy on her face.

Vera regarded Jane and looked over at Kal. “Well, then,” she sighed, “let’s get on with it.”

She sat back down at the scope and quickly located the fertilized egg. In a moment, she had injected what she hoped would be the magic bullet into each of the nuclei. She carried the dish over to an incubator, where it would sit for four or five days before the developing embryo would be transplanted into a recipient female cat.

Kal returned to the scope to look for more fertilized eggs. In the course of the next four hours, he and Vera found eighteen more and processed them in the same manner. Out of twenty-four oocytes, nineteen had become fertilized. Vera was not discouraged. She knew the success rate for IVF in cats was often lower.

For five days one or the other of the vets drove up to the institute to check on the developing embryos, to change the culture medium, and to inject the four future-recipient females with FSH. Vera was comforted by the fact that most of the eggs exhibited the expected cell division. Only four of the nineteen turned out to be non-viable. Meanwhile, Angelo sent daily e-mail messages relaying news from the CDC, as well as from France and Russia.

By Tuesday, three of the four recipient queens at the institute were showing signs of estrus. “Time to inject the gonadotropin,” said Vera. “Will you take care of that, Kal?”

“Sure. Looks like the embryos are about ready for transplanting. We have three new morulas today. I think we should start tomorrow and continue on Thursday, if necessary.”

“Tomorrow it is.”

Vera, Kal and Jane prepared for another day of surgery. With Noah watching from the observation room, the trio set up the equipment for next series of operations. Each of the queens would receive either three or four viable embryos.

Vera opened the door to the cage of the black-and-white female they had named Gertrude. The cat hissed briefly, but, as Vera stroked her back and plied her with catnip, she calmed down.

“That’s an appropriate name for a queen,” Jane remarked.

Vera nodded. “There, there, milady,” she said softly as she continued petting the animal. “We won’t hurt you much. Wouldn’t you like to be a savior of your species?”

The procedure for implanting the embryos was similar to that used to harvest the egg cells, but this time embryos were going in rather than eggs coming out. Vera manned the laparoscope, Kal again served as anesthesiologist, and Jane acted as surgical nurse. Vera deftly placed four embryos in Gertrude’s left uterine horn.

“Wouldn’t it be better to put two in one horn and two in the other?” asked Jane.

“You want to answer that, Kal?” Vera looked up from the cat at her colleague.

Kal grinned under his mask. “The reason is, Jane, that the embryos don’t get implanted right away. They move around quite a bit for several days, and some will migrate to the other horn. It’ll be several more days before the embryos finally settle down and become fixed to the uterine wall.”